Department of Biochemistry

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11-ketotestosterone and 11-ketodihydrotestosterone in castration resistant prostate cancer : potent androgens which can no longer be ignored
(Public Library of Science, 2016) Pretorius, Elzette; Africander, Donita J.; Vlok, Mare; Perkins, Meghan S.; Quanson, Jonathan; Storbeck, Karl-Heinz
Dihydrotestosterone (DHT) is regarded as the most potent natural androgen and is implicated in the development and progression of castration resistant prostate cancer (CRPC). Under castrate conditions, DHT is produced from the metabolism of the adrenal androgen precursors, DHEA and androstenedione. Recent studies have shown that the adrenal steroid 11β-hydroxyandrostenedione (11OHA4) serves as the precursor to the androgens 11-ketotestosterone (11KT) and 11-ketodihydrotestosterone (11KDHT). In this study we comprehensively assess the androgenic activity of 11KT and 11KDHT. This is the first study, to our knowledge, to show that 11KT and 11KDHT, like T and DHT, are potent and efficacious agonists of the human androgen receptor (AR) and induced both the expression of representative AR-regulated genes as well as cellular proliferation in the androgen dependent prostate cancer cell lines, LNCaP and VCaP. Proteomic analysis revealed that 11KDHT regulated the expression of more AR-regulated proteins than DHT in VCaP cells, while in vitro conversion assays showed that 11KT and 11KDHT are metabolized at a significantly lower rate in both LNCaP and VCaP cells when compared to T and DHT, respectively. Our findings show that 11KT and 11KDHT are bona fide androgens capable of inducing androgen-dependant gene expression and cell growth, and that these steroids have the potential to remain active longer than T and DHT due to the decreased rate at which they are metabolised. Collectively, our data demonstrates that 11KT and 11KDHT likely play a vital, but overlooked, role in the development and progression of CRPC.
11β-Hydroxyandrostenedione returns to the steroid arena : biosynthesis, metabolism and function
(MDPI, 2013-10) Bloem, Liezl M.; Storbeck, Karl-Heinz; Schloms, Lindie; Swart, Amanda C.
The biological significance of 11β-hydroxyandrostenedione (11OHA4) has eluded researchers for the past six decades. It is now known that 11OHA4 is biosynthesized in the androgen arm of the adrenal steroidogenesis pathway and subsequently metabolized by steroidogenic enzymes in vitro, serving as precursor to recognized and novel androgenic steroids. These in vitro findings extend beyond the adrenal, suggesting that 11OHA4 could be metabolized in steroid-responsive peripheral tissues, as is the case for androgen precursor metabolites of adrenal origin. The significance thereof becomes apparent when considering that the metabolism of 11OHA4 in LNCaP androgen dependent prostate cancer cells yields androgenic steroid metabolites. It is thus possible that 11OHA4 may be metabolized to yield ligands for steroid receptors in not only the prostate but also in other steroid-responsive tissues. Future investigations of 11OHA4 may therefore characterize it as a vital steroid with far-reaching physiological consequences. An overview of the research on 11OHA4 since its identification in 1953 will be presented, with specific focus on the most recent works that have advanced our understanding of its biological role, thereby underscoring its relevance in health and disease.
A fully dissociated compound of plant origin for inflammatory gene repression
(PNAS, 2005-11) De Bosscher, Karolien; Van den Berghe, Wim; Beck, Ilse M. E.; Van Molle, Wim; Hennuyer, Nathalie; Hapgood, Janet; Liber, Claude; Staels, Bart; Louw, Ann; Haegeman, Guy
The identification of selective glucocorticoid receptor (GR) modifiers, which separate transactivation and transrepression properties, represents an important research goal for steroid pharmacology. Although the gene-activating properties of GR are mainly associated with undesirable side effects, its negative interference with the activity of transcription factors, such as NF-κB, greatly contributes to its antiinflammatory and immune-suppressive capacities. In the present study, we found that Compound A (CpdA), a plant-derived phenyl aziridine precursor, although not belonging to the steroidal class of GR-binding ligands, does mediate gene-inhibitory effects by activating GR. We demonstrate that CpdA exerts an antiinflammatory potential by down-modulating TNF-induced proinflammatory gene expression, such as IL-6 and E-selectin, but, interestingly, does not at all enhance glucocorticoid response element-driven genes or induce GR binding to glucocorticoid response element-dependent genes in vivo. We further show that the specific gene-repressive effect of CpdA depends on the presence of functional GR, displaying a differential phosphorylation status with CpdA as compared with dexamethasone treatment. The antiinflammatory mechanism involves both a reduction of the in vivo DNA-binding activity of p65 as well as an interference with the transactivation potential of NF-κB. Finally, we present evidence that CpdA is as effective as dexamethasone in counter-acting acute inflammation in vivo and does not cause a hyperglycemic side effect. Taken together, this compound may be a lead compound of a class of antiinflammatory agents with fully dissociated properties and might thus hold great potential for therapeutic use. © 2005 by The National Academy of Sciences of the USA.
Abrogation of glucocorticoid receptor dimerization correlates with dissociated glucocorticoid behavior of compound A
(HighWire Press, 2010-03) Robertson, Steven; Allie-Reid, Fatima; Vanden Berghe, Wim; Visser, Koch; Binder, Anke; Africander, Donita; Vismer, Michael; De Bosscher, Karolien; Hapgood, Janet; Haegeman, Guy; Louw, Ann; A-7620-2012
Compound A (CpdA), a dissociated glucocorticoid receptor modulator, decreases corticosteroid-binding globulin (CBG), adrenocorticotropic hormone (ACTH), and luteneinizing hormone levels in rats. Whether this is due to transcriptional regulation by CpdA is not known. Using promoter reporter assays we show that CpdA, like dexamethasone (Dex), directly transrepresses these genes. Results using a rat Cbg proximal-promoter reporter construct in BWTG3 and HepG2 cell lines support a glucocorticoid receptor (GR)-dependent transrepression mechanism for CpdA. However, CpdA, unlike Dex, does not result in transactivation via glucocorticoid-responsive elements within a promoter reporter construct even when GR is co-transfected. The inability of CpdA to result in transactivation via glucocorticoid- responsive elements is confirmed on the endogenous tyrosine aminotransferase gene, whereas transrepression ability is confirmed on the endogenous CBG gene. Consistent with a role for CpdA in modulating GR activity, whole cell binding assays revealed that CpdA binds reversibly to the GR, but with lower affinity than Dex, and influences association of [3H]Dex, but has no effect on dissociation. In addition, like Dex, CpdA causes nuclear translocation of the GR, albeit to a lesser degree. Several lines of evidence, including fluorescence resonance energy transfer, co-immunoprecipitation, and nuclear immunofluorescence studies of nuclear localization- deficient GR show that CpdA, unlike Dex, does not elicit ligand-induced GR dimerization. Comparison of the behavior of CpdA in the presence of wild type GR to that of Dex with a dimerization-deficient GR mutant (GRdim) strongly supports the conclusion that loss of dimerization is responsible for the dissociated behavior of CpdA.
Acetyl-4′-phosphopantetheine is stable in serum and prevents phenotypes induced by pantothenate kinase deficiency
(Nature Research, 2017-09-12) Di Meo, Ivano; Colombelli, Cristina; Srinivasan, Balaji; De Villiers, Marianne; Hamada, Jeffrey; Jeong, Suh Y.; Fox, Rachel; Woltjer, Randall L.; Tepper, Pieter G.; Lahaye, Liza L.; Rizzetto, Emanuela; Harrs, Clara H.; De Boer, Theo; Van der Zwaag, Marianne; Jenko, Branko; Cusak, Alen; Pahor, Jerca; Kosec, Gregor; Grzeschik, Nicola A.; Hayflick, Susan J.; Tiranti, Valeria; Sibon, Ody C. M.
Coenzyme A is an essential metabolite known for its central role in over one hundred cellular metabolic reactions. In cells, Coenzyme A is synthesized de novo in five enzymatic steps with vitamin B5 as the starting metabolite, phosphorylated by pantothenate kinase. Mutations in the pantothenate kinase 2 gene cause a severe form of neurodegeneration for which no treatment is available. One therapeutic strategy is to generate Coenzyme A precursors downstream of the defective step in the pathway. Here we describe the synthesis, characteristics and in vivo rescue potential of the acetyl-Coenzyme A precursor S-acetyl-4′-phosphopantetheine as a possible treatment for neurodegeneration associated with pantothenate kinase deficiency.
Anti-inflammatory effects of Aspalathus linearis and Cyclopia spp. extracts in a UVB/keratinocyte (HaCaT) model utilising interleukin-1α accumulation as biomarker
(MDPI, 2016-10-02) Magcwebeba, Tandeka; Swart, Pieter; Swanevelder, Sonja; Joubert, Elizabeth; Gelderblom, Wentzel
Ultraviolet B (UVB) radiation is one of the major predisposing risk factors of skin cancer. The anticancer and photoprotective effects of unoxidized rooibos (Aspalathus linearis) and honeybush (Cyclopia) herbal teas, containing high levels of dihydrochalones and xanthones, respectively, have been demonstrated in skin cancer models in vivo. In the current study, the anti-inflammatory effects of methanol and aqueous extracts of these herbal teas were investigated in a UVB/HaCaT keratinocyte model with intracellular interleukin-1α (icIL-1α) accumulation as a biomarker. Extracts of green tea (Camellia sinensis) served as benchmark. Both extracts of green tea and rooibos, as well as the aqueous extract of C. intermedia, enhanced UVB-induced inhibition of cell viability, proliferation and induction of apoptosis, facilitating the removal of icIL-1α. The underlying mechanisms may involve mitochondrial dysfunction exhibiting pro-oxidant responses via polyphenol-iron interactions. The methanol extracts of honeybush, however, protected against UVB-induced reduction of cell growth parameters, presumably via antioxidant mechanisms that prevented the removal of highly inflamed icIL-1α-containing keratinocytes via apoptosis. The dual antioxidant and/or pro-oxidant role of the polyphenolic herbal tea constituents should be considered in developing preventive strategies against UVB-induced skin carcinogenesis. The indirect removal of UVB damaged keratinocytes by herbal tea extracts via apoptosis may find application in the prevention of photo-induced inflammation.
Antimikrobiese nanovesels vir waterbehandeling : poli(vinielalkohol)- en poli(akrielonitriel)- nanovesels met silwer-nanopartikels
(LitNet, 2012) Du Plessis, Danielle; Botes, Marelize; Dicks, Leon Milner Theodore, 1961-; Cloete, Thomas Eugene
Daar moet verbeter word op bestaande watersuiweringsmetodes ten einde mikrobiologies veilige en bekostigbare drinkwater te verskaf. Nanovesels word reeds gebruik in waterfiltrasiesisteme en nanofiltrasie mag selfs as ’n alternatief vir biosiede gebruik word. Verskeie variasies van nanovesels met biosiede is in die onlangse literatuur omskryf. Omdat nanovesels met antimikrobiese aktiwiteit ’n relatief nuwe studieveld is, is nog weinig studies gewy aan die uiteensetting van praktiese standaardmetodes vir antimikrobiese-aktiwiteit-bepaling. Die aktiwiteit van antimikrobiese vesels word oor die algemeen met plaattellings van kolonievormende eenhede (KVE) bepaal. Hierdie metode bepaal ’n afname in die getal kweekbare patogeniese selle teenwoordig. Die hoofdoel van hierdie studie was om ’n vinnige, maklik uitvoerbare en akkurate toets te ontwikkel om die aktiwiteit van antimikrobiese nanovesels te bepaal.
The biodiversity hotspot as evolutionary hot-bed : spectacular radiation of Erica in the Cape Floristic Region
(BioMed Central, 2016-09-17) Pirie, M. D.; Oliver, E. G. H.; De Kuppler, A. Mugrabi; Gehrke, B.; Le Maitre, N. C.; Kandziora, M.; Bellstedt, D. U.
Background: The disproportionate species richness of the world’s biodiversity hotspots could be explained by low extinction (the evolutionary “museum”) and/or high speciation (the “hot-bed”) models. We test these models using the largest of the species rich plant groups that characterise the botanically diverse Cape Floristic Region (CFR): the genus Erica L. We generate a novel phylogenetic hypothesis informed by nuclear and plastid DNA sequences of c. 60 % of the c. 800 Erica species (of which 690 are endemic to the CFR), and use this to estimate clade ages (using RELTIME; BEAST), net diversification rates (GEIGER), and shifts in rates of diversification in different areas (BAMM; MuSSE). Results: The diversity of Erica species in the CFR is the result of a single radiation within the last c. 15 million years. Compared to ancestral lineages in the Palearctic, the rate of speciation accelerated across Africa and Madagascar, with a further burst of speciation within the CFR that also exceeds the net diversification rates of other Cape clades. Conclusions: Erica exemplifies the “hotbed” model of assemblage through recent speciation, implying that with the advent of the modern Cape a multitude of new niches opened and were successively occupied through local species diversification.
Blood ketone bodies and breath acetone analysis and their correlations in type 2 diabetes mellitus
(MDPI, 2019-12-17) Saasa, Valentine; Beukes, Mervyn; Lemmer, Yolandy; Mwakikunga, Bonex
Analysis of volatile organic compounds in the breath for disease detection and monitoring has gained momentum and clinical significance due to its rapid test results and non-invasiveness, especially for diabetes mellitus (DM). Studies have suggested that breath gases, including acetone, may be related to simultaneous blood glucose (BG) and blood ketone levels in adults with types 2 and 1 diabetes. Detecting altered concentrations of ketones in the breath, blood and urine may be crucial for the diagnosis and monitoring of diabetes mellitus. This study assesses the efficacy of a simple breath test as a non-invasive means of diabetes monitoring in adults with type 2 diabetes mellitus. Human breath samples were collected in Tedlar™ bags and analyzed by headspace solid-phase microextraction and gas chromatography-mass spectrometry (HS-SPME/GC-MS). The measurements were compared with capillary BG and blood ketone levels (β-hydroxybutyrate and acetoacetate) taken at the same time on a single visit to a routine hospital clinic in 30 subjects with type 2 diabetes and 28 control volunteers. Ketone bodies of diabetic subjects showed a significant increase when compared to the control subjects; however, the ketone levels were was controlled in both diabetic and non-diabetic volunteers. Worthy of note, a statistically significant relationship was found between breath acetone and blood acetoacetate (R = 0.89) and between breath acetone and β-hydroxybutyrate (R = 0.82).
The carbon switch at the level of pyruvate and phosphoenolpyruvate in sulfolobus solfataricus P2
(Frontiers Media, 2019-04-12) Haferkamp, Patrick; Tjaden, Britta; Shen, Lu; Brasen, Christopher; Kouril, Theresa; Siebers, Bettina
Sulfolobus solfataricus P2 grows on different carbohydrates as well as alcohols, peptides and amino acids. Carbohydrates such as D-glucose or D-galactose are degraded via the modified, branched Entner–Doudoroff (ED) pathway whereas growth on peptides requires the Embden–Meyerhof–Parnas (EMP) pathway for gluconeogenesis. As for most hyperthermophilic Archaea an important control point is established at the level of triosephophate conversion, however, the regulation at the level of pyruvate/phosphoenolpyruvate conversion was not tackled so far. Here we describe the cloning, expression, purification and characterization of the pyruvate kinase (PK, SSO0981) and the phosphoenolpyruvate synthetase (PEPS, SSO0883) of Sul. solfataricus. The PK showed only catabolic activity [catalytic efficiency (PEP): 627.95 mM⁻¹s⁻¹, 70°C] with phosphoenolpyruvate as substrate and ADP as phosphate acceptor and was allosterically inhibited by ATP and isocitrate (Ki 0.8 mM). The PEPS was reversible, however, exhibited preferred activity in the gluconeogenic direction [catalytic efficiency (pyruvate): 1.04 mM⁻¹s⁻¹, 70°C] and showed some inhibition by AMP and α-ketoglutarate. The gene SSO2829 annotated as PEPS/pyruvate:phosphate dikinase (PPDK) revealed neither PEPS nor PPDK activity. Our studies suggest that the energy charge of the cell as well as the availability of building blocks in the citric acid cycle and the carbon/nitrogen balance plays a major role in the Sul. solfataricus carbon switch. The comparison of regulatory features of well-studied hyperthermophilic Archaea reveals a close link and sophisticated coordination between the respective sugar kinases and the kinetic and regulatory properties of the enzymes at the level of PEP-pyruvate conversion.
Chemoprevention of LA7-induced mammary tumor growth by SM6Met, a well-characterized cyclopia extract
(Frontiers, 2018) Oyenihi, Omolola R.; Krygsman, Annadie; Verhoog, Nicolette; De Beer, Dalene; Saayman, Michael J.; Mouton, Thys M.; Louw, Ann; Wong, Vincent Kam Wai
Breast cancer (BC) is the leading cause of cancer-related deaths in women. Chemoprevention of BC by using plant extracts is gaining attention. SM6Met, a wellcharacterized extract of Cyclopia subternata with reported selective estrogen receptor subtype activity, has shown tumor suppressive effects in a chemically induced BC model in rats, which is known to be estrogen responsive. However, there is no information on the estrogen sensitivity of the relatively new orthotopic model of LA7 cell-induced mammary tumors. In the present study, the potential chemopreventative and side-effect profile of SM6Met on LA7 cell-induced tumor growth was evaluated, as was the effects of 17b-estradiol and standard-of-care (SOC) endocrine therapies, such as tamoxifen (TAM), letrozole (LET), and fulvestrant (FUL). Tumor growth was observed in the tumorvehicle control group until day 10 post tumor induction, which declined afterward on days 12–14. SM6Met suppressed tumor growth to the same extent as TAM, while LET, but not FUL, also showed substantial anti-tumor effects. Short-term 17b-estradiol treatment reduced tumor volume on days prior to day 10, whereas tumor promoting effects were observed during long-term treatment, which was especially evident at later time points. Marked elevation in serum markers of liver injury, which was further supported by histological evaluation, was observed in the vehicle-treated tumor control, TAM, LET, and long-term 17b-estradiol treatment groups. Alterations in the lipid profiles were also observed in the 17b-estradiol treatment groups. In contrast, SM6Met did not augment the increase in serum levels of liver injury biomarkers caused by tumor induction and no effect was observed on lipid profiles. In summary, the results from the current study demonstrate the chemopreventative effect of SM6Met on mammary tumor growth, which was comparable to that of TAM, without eliciting the negative side-effects observed with this SOC endocrine therapy. Furthermore, the results of this study also showed some responsiveness of LA7-induced tumors to estrogen and SOC endocrine therapies. Thus, this model may be useful in evaluating potential endocrine therapies for hormone responsive BC. Keywords: chemoprevention, Cyclopia, mammary tumor, phytoestrogen, tamoxifen, letrozole, fulvestrant
Comparative characterization of plasmodium falciparum Hsp70-1 relative to E. coli DnaK reveals the functional specificity of the parasite chaperone
(Multidisciplinary Digital Publishing Institute (MDPI), 2020-06-04) Lebepe, Charity Mekgwa; Matambanadzo, Pearl Rutendo; Makhoba, Xolani Henry; Achilonu, Ikechukwu; Zininga, Tawanda; Shonhai, Addmore
Hsp70 is a conserved molecular chaperone. How Hsp70 exhibits specialized functions across species remains to be understood. Plasmodium falciparum Hsp70-1 (PfHsp70-1) and Escherichia coli DnaK are cytosol localized molecular chaperones that are important for the survival of these two organisms. In the current study, we investigated comparative structure-function features of PfHsp70-1 relative to DnaK and a chimeric protein, KPf, constituted by the ATPase domain of DnaK and the substrate binding domain (SBD) of PfHsp70-1. Recombinant forms of the three Hsp70s exhibited similar secondary and tertiary structural folds. However, compared to DnaK, both KPf and PfHsp70-1 were more stable to heat stress and exhibited higher basal ATPase activity. In addition, PfHsp70-1 preferentially bound to asparagine rich peptide substrates, as opposed to DnaK. Recombinant P. falciparum adenosylmethionine decarboxylase (PfAdoMetDC) co-expressed in E. coli with either KPf or PfHsp70-1 was produced as a fully folded product. Co-expression of PfAdoMetDC with heterologous DnaK in E. coli did not promote folding of the former. However, a combination of supplementary GroEL plus DnaK improved folding of PfAdoMetDC. These findings demonstrated that the SBD of PfHsp70-1 regulates several functional features of the protein and that this molecular chaperone is tailored to facilitate folding of plasmodial proteins.
Complex stability and dynamic subunit interchange modulates the disparate activities of the yeast moonlighting proteins Hal3 and Vhs3
(Springer Nature, 2015-10) Abrie, J. Albert; Molero, Cristina; Arino, Joaquin; Strauss, Erick
Saccharomyces cerevisiae Hal3 and Vhs3 are moonlighting proteins, acting both as inhibitors of the serine/threonine protein phosphatase Ppz1 and as subunits (together with Cab3) of the unique heterotrimeric phosphopantothenoylcysteine decarboxylase (PPCDC) enzyme of Hemiascomycetous yeast. Both these roles are essential: PPCDC catalyses the third step of coenzyme A biosynthesis, while Ppz1 inhibition is required for regulation of monovalent cation homeostasis. However, the mechanisms by which these proteins’ disparate activities are regulated are not well understood. The PPCDC domains (PDs) of Hal3, Vhs3 and Cab3 constitute the minimum requirement for these proteins to show both PPCDC activity and, in the case of Hal3 and Vhs3, to bind to Ppz1. Using these PD proteins as a model system to study the possibility of dynamic interchange between these roles, we provide evidence that Hal3 binds Ppz1 as a monomer (1:1 stoichiometry), requiring it to deoligomerize from its usual homo- and heterotrimeric states (the latter having PPCDC activity). This de-oligomerization is made possible by structural features that set Hal3 apart from Vhs3, increasing its ability to undergo monomer exchange. These findings suggest that oligomer interchange may be a significant factor in the functional regulation of these proteins and their various unrelated (moonlighting) functions.
Conservation implications of avian malaria exposure for African penguins during rehabilitation
(Academy of Science of South Africa, 2017-02-15) Botes, Annelise; Thiart, Hanlie; Parsons, Nola J.; Bellstedt, Dirk U.
ENGLISH ABSTRACT: The African penguin (Spheniscus demersus) is the only penguin species that breeds on the African continent and it is currently classified as endangered. Its conservation is assisted by the Southern African Foundation for the Conservation of Coastal Birds (SANCCOB) which is a seabird rehabilitation facility based at the Rietvlei Wetland Reserve in Tableview, Cape Town. Despite the success of SANCCOB in rehabilitating diseased, injured or oiled penguins, significant mortalities have occurred at the facility as a result of avian malaria. Avian malaria can be contracted during rehabilitation during which penguins are inadvertently exposed to additional threats. An enzyme-linked immunosorbent assay (ELISA) was used to assess the anti-Plasmodium antibody levels of penguins to avian malaria on entry into the SANCCOB facility from 2001 to 2004 and during their rehabilitation process. Using blood smear data, avian malaria prevalence and malaria-related deaths were also monitored from 2002 to 2013. Significant increases in anti-Plasmodium antibody levels after admission were found during summer months. New infection and not parasite recrudescence was concluded to be the cause of this increase. This source was confirmed by a dramatic drop in penguin mortalities upon exclusion of mosquito vectors in 2008. Mortalities did not depend on the birds’ abilities to produce an anti-Plasmodium antibody response and oiling had no influence on immunity or prevalence of avian malaria infections. This study highlights the importance of mosquito vector control to control pathogen exposure in wild bird rehabilitation centres. Significance: • Efforts to assist with the conservation of endangered species can unintentionally add to the conservation burden. • Rehabilitation influences exposure of African penguins to avian malaria. • Avian malaria prevalence and mortality are not influenced by oiling or anti-Plasmodium antibody responses. • Vector control can limit avian malaria exposure in wild bird rehabilitation centres.
Consistent phenological shifts in the making of a biodiversity hotspot : the Cape flora
(BioMed Central, 2011-02) Warren, Ben H.; Bakker, Freek T.; Bellstedt, Dirk U.; Bytebier, Benny; Claszen-Bockhoff, Regine; Dreyer, Leanne L.; Edwards, Dawn; Forest, Felix; Galley, Chloe; Hardy, Christopher R.; Linder, H. Peter; Muasya, A. Muthama; Mummenhoff, Klaus; Oberlander, Kenneth C.; Quint, Marcus; Richardson, James E.; Savolainen, Vincent; Schrire, Brian D.; Van der Niet, Timotheus; Verboom, G. Anthony; Yesson, Christopher; Hawkins, Julie A.
ABSTRACT: Background: The best documented survival responses of organisms to past climate change on short (glacial-interglacial) timescales are distributional shifts. Despite ample evidence on such timescales for local adaptations of populations at specific sites, the long-term impacts of such changes on evolutionary significant units in response to past climatic change have been little documented. Here we use phylogenies to reconstruct changes in distribution and flowering ecology of the Cape flora - South Africa's biodiversity hotspot - through a period of past (Neogene and Quaternary) changes in the seasonality of rainfall over a timescale of several million years. Results: Forty-three distributional and phenological shifts consistent with past climatic change occur across the flora, and a comparable number of clades underwent adaptive changes in their flowering phenology (9 clades; half of the clades investigated) as underwent distributional shifts (12 clades; two thirds of the clades investigated). Of extant Cape angiosperm species, 14-41% have been contributed by lineages that show distributional shifts consistent with past climate change, yet a similar proportion (14-55%) arose from lineages that shifted flowering phenology. Conclusions: Adaptive changes in ecology at the scale we uncover in the Cape and consistent with past climatic change have not been documented for other floras. Shifts in climate tolerance appear to have been more important in this flora than is currently appreciated, and lineages that underwent such shifts went on to contribute a high proportion of the flora's extant species diversity. That shifts in phenology, on an evolutionary timescale and on such a scale, have not yet been detected for other floras is likely a result of the method used; shifts in flowering phenology cannot be detected in the fossil record.
Cyclopia extracts act as ERα antagonists and ERβ agonists, in vitro and in vivo
(Public Library of Science (PLOS), 2013-11) Visser, Koch; Mortimer, Morne; Louw, Ann
Hormone replacement therapy associated risks, and the concomitant reluctance of usage, has instigated the search for new generations of estrogen analogues that would maintain estrogen benefits without associated risks. Furthermore, if these analogues display chemo-preventative properties in breast and endometrial tissues it would be of great value. Both the selective estrogen receptor modulators as well as the selective estrogen receptor subtype modulators have been proposed as estrogen analogues with improved risk profiles. Phytoestrogen containing extracts of Cyclopia, an indigenous South African fynbos plant used to prepare Honeybush tea may serve as a source of new estrogen analogues. In this study three extracts, P104, SM6Met, and cup-of-tea, from two species of Cyclopia, C. genistoides and C. subternata, were evaluated for ER subtype specific agonism and antagonism both in transactivation and transrepression. For transactivation, the Cyclopia extracts displayed ERα antagonism and ERβ agonism when ER subtypes were expressed separately, however, when co-expressed only agonism was uniformly observed. In contrast, for transrepression, this uniform behavior was lost, with some extracts (P104) displaying uniform agonism, while others (SM6Met) displayed antagonism when subtypes were expressed separately and agonism when co-expressed. In addition, breast cancer cell proliferation assays indicate that extracts antagonize cell proliferation in the presence of estrogen at lower concentrations than that required for proliferation. Furthermore, lack of uterine growth and delayed vaginal opening in an immature rat uterotrophic model validates the ERα antagonism of extracts observed in vitro and supports the potential of the Cyclopia extracts as a source of estrogen analogues with a reduced risk profile.
Data comparing the separation and elution of vitamin D metabolites on an ultra performance supercritical fluid chromatography tandem-mass spectrometer (UPSFC-MS/MS) compared to liquid chromatography (LC) and data presenting approaches to UPSFC method optimization
(Elsevier, 2018) Jenkinson, Carl; Taylor, Angela E.; Storbeck, Karl-Heinz; Hewison, Martin
The data presented is related to the research article "Analysis of multiple vitamin D metabolites by ultra performance supercritical fluid chromatography-tandem mass spectrometry (UPSFC-MS/MS)" (Jenkinson et al., 2018) [1]. This article will include data obtained from method development, optimization and analysis of multiple vitamin D metabolites on an ultra performance supercritical fluid chromatography tandem-mass spectrometry (UPSFC-MS/MS). This includes chromatograms from column screening to confirm the most suitable column for analyte separation. Additionally, further chromatograms and figures compare separation and analyte signal strength during the optimization of other UPSFC parameters. Mass spectra will demonstrate the optimization of MS conditions for the UPSFC-MS/MS method. Chromatogram data from UHPLC vitamin D analysis is also presented in order to compare the separation and elution of vitamin D metabolites using UPSFC and UHPLC. This data will highlight the outputs that aid in method development and identifying the separation technique suited for vitamin D quantitation.
Degradation of proteins and starch by combined immobilization of protease, α-amylase and β-galactosidase on a single electrospun nanofibrous membrane
(MDPI, 2019-01-31) Cloete, William J.; Hayward, Stefan; Swart, Pieter; Klumperman, Bert
Two commercially available enzymes, Dextrozyme (α-amylase) and Esperase (protease), were covalently immobilized on non-woven electrospun poly(styrene-co-maleic anhydride) nanofiber mats with partial retention of their catalytic activity. Immobilization was achieved for the enzymes on their own as well as in different combinations with an additional enzyme, β-galactosidase, on the same non-woven nanofiber mat. This experiment yielded a universal method for immobilizing different combinations of enzymes with nanofibrous mats containing maleic anhydride (MAnh) residues in the polymer backbone.
Degradation of synthetic xylan effluent using a membrane bioreactor
(Academy of Science for South Africa, 2003) Edward, V. A.; Pillay, V. L.; Swart, P.; Jacobs, E.; Singh, S.
We have built a novel membrane bioreactor for the degradation of synthetic xylan effluent. The reactor contains 30 internally skinned polysulphone membranes as an immobilization matrix for xylanase, the degrading agent, and was constructed with stainless steel to withstand high temperatures, as Thermomyces lanuginosus SSBP xylanase has an optimum temperature of 50°C. Overall, 85.1% of the xylanase was immobilized onto the polysulphone membranes by adsorption. Preliminary results showed that the immobilized enzyme was capable of degrading the xylan effluent. Prior to contact with xylanase, there was 0.0 μg ml-1 xylose, 14.2 μg ml-1 xylobiose and 7.2 μg ml-1 xylotriose present in the xylan effluent. After 180 min, the xylose, xylobiose and xylotriose concentrations were 246, 103 and 91 μg ml-1, respectively. Substantial increase in degradation products is promising for the development of a larger-scale bioreactor for effluent treatment.
Delving deeper : relating the behaviour of a metabolic system to the properties of its components using symbolic metabolic control analysis
(Public Library of Science, 2018-11-28) Christensen, Carl D.; Hofmeyr, Jan-Hendrik S.; Rohwer, Johann M.
High-level behaviour of metabolic systems results from the properties of, and interactions between, numerous molecular components. Reaching a complete understanding of metabolic behaviour based on the system’s components is therefore a difficult task. This problem can be tackled by constructing and subsequently analysing kinetic models of metabolic pathways since such models aim to capture all the relevant properties of the system components and their interactions. Symbolic control analysis is a framework for analysing pathway models in order to reach a mechanistic understanding of their behaviour. By providing algebraic expressions for the sensitivities of system properties, such as metabolic flux or steadystate concentrations, in terms of the properties of individual reactions it allows one to trace the high level behaviour back to these low level components. Here we apply this method to a model of pyruvate branch metabolism in Lactococcus lactis in order to explain a previously observed negative flux response towards an increase in substrate concentration. With this method we are able to show, first, that the sensitivity of flux towards changes in reaction rates (represented by flux control coefficients) is determined by the individual metabolic branches of the pathway, and second, how the sensitivities of individual reaction rates towards their substrates (represented by elasticity coefficients) contribute to this flux control. We also quantify the contributions of enzyme binding and mass-action to enzyme elasticity separately, which allows for an even finer-grained understanding of flux control. These analytical tools allow us to analyse the control properties of a metabolic model and to arrive at a mechanistic understanding of the quantitative contributions of each of the enzymes to this control. Our analysis provides an example of the descriptive power of the general principles of symbolic control analysis.

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