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1.  Bioprospecting Keratinous Materials 
The concept of bioprospecting for bioactive peptides from keratin-containing materials such as wool, hair, skin and feathers presents an exciting opportunity for discovery of novel functional food ingredients and nutraceuticals, while value-adding to cheap and plentiful natural sources. The published literature reports multiple examples of proline-rich peptides with productive bio-activity in models of human disease including tumour formation, hypertension control and Alzheimer’s disease. Bioactive peptides have been identified from food and other protein sources however the bioactivity of keratin-related proteins and peptides is largely unknown. Considering the high representation of proline-rich peptides among proven bioactive peptides, the proline-rich character of keratinous proteins supports current research. A selection of mammalian (cow epidermis, sheep wool) and avian (chicken feather) keratinous materials were subjected to enzymatic hydrolysis using established processing methods. A bio-assay of determining inhibition of early stage amyloid aggregation involved using a model fibril-forming protein – reduced and carboxymethylated bovine K-casein (RCMk-CN) and quantitation of fibril development with the amyloid-specific fluorophore, Thioflavin T (ThT). The assay was fully validated for analytical repeatability and used together with appropriate positive controls. Peptide library products derived from chicken feather (n=9), sheep wool (n=9) and bovine epidermis (n=9) were screened in the fibril inhibition assay based on K-casein. 3 of 27 products exhibited interesting levels of bio-activity with regard to fibril inhibition. HPLC profiles provide an indication of the complexity of the assemblage of peptides in the three active products. We conclude the bioprospecting research using keratinous materials shows promise for discovery of useful bioactive peptides.
PMCID: PMC3002413  PMID: 21188026
Bioactive peptides; bioprospecting; keratin
2.  The genome of the simian and human malaria parasite Plasmodium knowlesi 
Nature  2008;455(7214):799-803.
Plasmodium knowlesi is an intracellular malaria parasite whose natural vertebrate host is Macaca fascicularis (the ‘kra’ monkey); however, it is now increasingly recognized as a significant cause of human malaria, particularly in southeast Asia1,2. Plasmodium knowlesi was the first malaria parasite species in which antigenic variation was demonstrated3, and it has a close phylogenetic relationship to Plasmodium vivax​4, the second most important species of human malaria parasite (reviewed in ref. 4). Despite their relatedness, there are important phenotypic differences between them, such as host blood cell preference, absence of a dormant liver stage or ‘hypnozoite’ in P. knowlesi, and length of the asexual cycle (reviewed in ref. 4). Here we present an analysis of the P. knowlesi (H strain, Pk1(A+) clone5) nuclear genome sequence. This is the first monkey malaria parasite genome to be described, and it provides an opportunity for comparison with the recently completed P. vivax genome4 and other sequenced Plasmodium genomes6-8. In contrast to other Plasmodium genomes, putative variant antigen families are dispersed throughout the genome and are associated with intrachromosomal telomere repeats. One of these families, the KIRs9, contains sequences that collectively match over one-half of the host CD99 extracellular domain, which may represent an unusual form of molecular mimicry.
PMCID: PMC2656934  PMID: 18843368
3.  Structure/function studies of dogfish α-crystallin, comparison with bovine α-crystallin 
Molecular Vision  2009;15:2411-2420.
α-Crystallin is the major protein of the mammalian lens where it contributes to the refractive properties needed for vision and possibly to the stability of the tissue. The aim of this study was to determine whether the properties of α-crystallin have changed during the course of evolution.
Dogfish α-crystallin, which appeared over 420 million years ago, has been contrasted with bovine α-crystallin, which emerged around 160 million years later, by comparing their sizes, the microenvironments of their cysteine and tryptophan residues, their chaperone-like activities and the flexibility of their COOH-terminal extensions.
Dogfish α-crystallin consists of αA- and αB-polypeptides, in a 1:5 ratio, and has a molecular mass of around 400 kDa. By contrast, the bovine protein is around 600-800 kDa in mass and has a 3:1 subunit ratio. Cysteine residues in the proteins were equally accessible to reaction with 5,5'-dithiobis-(2-nitrobenzoic acid). Quenching of fluorescence with acrylamide indicated tryptophan residues in the two proteins were in similar environments. The chaperone activity of dogfish α-crystallin was comparable to that of bovine α-crystallin in preventing the heat-induced precipitation of βL-crystallin but the dogfish protein was three times more effective at preventing insulin precipitation after reduction at 37 ˚C. 1H nuclear magnetic resonance spectroscopic studies showed that the last 17 amino acids of the dogfish αB polypeptide (V162-K178) have great conformational flexibility, are highly exposed to solvent and adopt little ordered conformation. This is comparable to, but slightly longer in length, than the COOH-terminal extension observed in mammalian α-crystallins.
The structure and properties of α-crystallin have changed relatively little during the evolutionary period from the emergence of sharks and mammals.
PMCID: PMC2785718  PMID: 19956560
4.  Human immunodeficiency virus type 1 membrane fusion mediated by a laboratory-adapted strain and a primary isolate analyzed by resonance energy transfer. 
Journal of Virology  1996;70(9):6437-6441.
Previous studies of human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein-mediated membrane fusion have focused on laboratory-adapted T-lymphotropic strains of the virus. The goal of this study was to characterize membrane fusion mediated by a primary HIV-1 isolate in comparison with a laboratory-adapted strain. To this end, a new fusion assay was developed on the basis of the principle of resonance energy transfer, using HeLa cells stably transfected with gp120/gp41 from the T-lymphotropic isolate HIV-1LA1 or the macrophage-tropic primary isolate HIV-1JR-FL. These cells fused with CD4+ target cell lines with a tropism mirroring that of infection by the two viruses. Of particular note, HeLa cells expressing HIV-1JR-FL gp120/gp41 fused only with PM1 cells, a clonal derivative of HUT 78, and not with other T-cell or macrophage cell lines. These results demonstrate that the envelope glycoproteins of these strains play a major role in mediating viral tropism. Despite significant differences exhibited by HIV-1JR-FL and HIV-1LAI in terms of tropism and sensitivity to neutralization by CD4-based proteins, the present study found that membrane fusion mediated by the envelope glycoproteins of these viruses had remarkably similar properties. In particular, the degree and kinetics of membrane fusion were similar, fusion occurred at neutral pH and was dependent on the presence of divalent cations. Inhibition of HIV-1JR-FL envelope glycoprotein-mediated membrane fusion by soluble CD4 and CD4-IgG2 occurred at concentrations similar to those required to neutralize this virus. Interestingly, higher concentrations of these agents were required to inhibit HIV-1LAI envelope glycoprotein-mediated membrane fusion, in contrast to the greater sensitivity of HIV-1LAI virions to neutralization by soluble CD4 and CD4-IgG2. This finding suggests that the mechanisms of fusion inhibition and neutralization of HIV-1 are distinct.
PMCID: PMC190675  PMID: 8709277
6.  Epstein's Brain 
British Medical Journal  1959;1(5124):791.
PMCID: PMC1992950

Results 1-8 (8)