Amyloids are filamentous protein deposits ranging in size from nanometres to microns and composed of aggregated peptide beta-sheets formed from parallel or anti-parallel alignments of peptide beta-strands. Amyloid-forming proteins have attracted a great deal of recent attention because of their association with over 30 diseases, notably neurodegenerative conditions like Alzheimer's, Huntington's, Parkinson's, Creutzfeldt-Jacob and prion disorders, but also systemic diseases such as amyotrophic lateral sclerosis (Lou Gehrig's disease) and type II diabetes. These diseases are all thought to involve important conformational changes in proteins, sometimes termed misfolding, that usually produce beta-sheet structures with a strong tendency to aggregate into water-insoluble fibrous polymers. Reasons for such conformational changes in vivo are still unclear. Intermediate aggregated state(s), rather than precipitated insoluble polymeric aggregates, have recently been implicated in cellular toxicity and may be the source of aberrant pathology in amyloid diseases. Numerous in vitro studies of short and medium length peptides that form amyloids have provided some clues to amyloid formation, with an alpha-helix to beta-sheet folding transition sometimes implicated as an intermediary step leading to amyloid formation. More recently, quite a few non-pathological amyloidogenic proteins have also been identified and physiological properties have been ascribed, challenging previous implications that amyloids were always disease causing. This article summarises a great deal of current knowledge on the occurrence, structure, folding pathways, chemistry and biology associated with amyloidogenic peptides and proteins and highlights some key factors that have been found to influence amyloidogenesis.

One of the important determinants of virulence of Mycobacterium tuberculosis is adaptation to adverse conditions encountered in the host cells. The ability of Mycobacterium to successfully adapt to stress conditions is brought about by the expression of specific regulons effected by a repertoire of σ factors. The induction and availability of σ factors in response to specific stimuli is governed by a complex regulatory network comprising a number of proteins, including σ factors themselves. A serine–threonine protein kinase‐mediated signaling pathway adds another dimension to the mycobacterial σ factor regulatory network. This review highlights the recent advances in understanding mycobacterial σ factors, their regulation and contribution to bacterial pathogenesis.

Lethal toxin from Bacillus anthracisis composed of protective antigen (PA) and lethal factor (LF). Anti-PA mAbs that neutralized lethal toxin activity, either in vivo or in vitro, identified three non-overlapping antigenic regions on PA. Two distinct antigenic regions were recognized by the four mAbs that neutralized lethal toxin activity by inhibiting the binding of 125I-LF to cell-bound PA. Mapping showed that one mAb, 1G3PA63, recognized an epitope on a 17 IcDa fragment located between amino acid residues Ser-168 and Phe-314. The three other mAbs, 2D3PA, 2D5PA and 10D2PA, recognized an epitope between amino acids Ile-581 and Asn-601. A single antigenic region was recognized by the three mAbs, 3B6PA, 14B7PA and 10E10PA63, that inhibited binding of 125I-PA to cells. This region was located between amino acids Asp-671 and lle-721. These results confirm previously defined functional domains of PA and suggest that LF may interact with two different sites on PA to form lethal toxin.

SummaryProtein tyrosine kinases and tyrosine phosphatases from several bacterial pathogens have been shown to act as virulence factors by modulating the phosphorylation and dephosphorylation of host proteins. The identification and characterization of two tyrosine phosphatases namely MptpA and MptpB from Mycobacterium tuberculosis has been reported earlier.MptpB is secreted by M. tuberculosis into extracellular mileu and exhibits a pH optimum of 5.6, similar to the pH of the lysosomal compartment of the cell. To determine the role of MptpB in the pathogenesis of M. tuberculosis , we constructed a mptpB mutant strain by homologous recombination and compared the ability of parent and the mutant strain to survive intracellularly. We show that disruption of the mptpB gene impairs the ability of the mutant strain to survive in activated macrophages and guinea pigs but not in resting macrophages suggesting the importance of its role in the host-pathogen interaction. Infection of guinea pigs with the mutant strain resulted in a 70-fold reduction in the bacillary load of spleens in infected animals as compared with the bacillary load in animals infected with the parental strain. Upon reintroduction of the mptpB gene into the mutant strain, the complemented strain was able to establish infection and survive in guinea pigs at rates comparable to the parental strain. These observations demonstrate a role of MptpB in the pathogenesis of M. tuberculosis .

A panel of variants with alanine substitutions in the small loop of anthrax toxin protective antigen domain 4 was created to determine individual amino acid residues critical for interactions with the cellular receptor and with a neutralizing monoclonal antibody, 14B7. Substituted protective antigen proteins were analyzed by cellular cytotoxicity assays, and their interactions with antibody were measured by plasmon surface resonance and analytical ultracentrifugation. Residue Asp 683 was the most critical for cell binding and toxicity, causing an ϳ1000-fold reduction in toxicity, but was not a large factor for interactions with 14B7. Substitutions in residues Tyr 681 , Asn 682 , and Pro 686 also reduced toxicity significantly, by 10 -100-fold. Of these, only Asn 682 and Pro 686 were also critical for interactions with 14B7. However, residues Lys 684 , Leu 685 , Leu 687 , and Tyr 688 were critical for 14B7 binding without greatly affecting toxicity. The K684A and L685A variants exhibited wild type levels of toxicity in cell culture assays; the L687A and Y688A variants were reduced only 1.5-and 5-fold, respectively.Bacillus anthracis secretes two toxins: edema toxin and lethal toxin. Each is composed of a common binding component, protective antigen (PA), 1 together with an enzymatic component, edema factor (EF), in the case of edema toxin and lethal factor (LF) in the case of lethal toxin (1-3). The current model for toxin entry into the cell illustrates the centrality of PA for toxin action. PA binds to cellular receptors, recently identified as splice variants of either tumor endothelial marker 8 (TEM8) (4 -6) or the closely related capillary morphogenesis protein 2 (CMG2) (7). Furin cleaves PA, releasing a 20-kDa fragment and leaving behind a 63-kDa portion (PA 63 ) capable of forming a heptamer, which has a newly exposed surface that binds . Heptamer complexes enter the endocytic pathway by receptor-mediated endocytosis (13), and upon acidification of the vesicle, the PA 63 heptamer undergoes a conformational change to form a pore through which EF and LF translocate into the cytoplasm (10, 11, 14 -16). Once in the cytoplasm, EF and LF exert their toxic effects.The PA protein can be divided into four domains based on its crystal structure, and functions can be attributed to the different domains based on mutational and biochemical analyses (16). Domain 1 (residues 1-258) contains the furin cleavage site as well as the hydrophobic portion of PA, which is exposed upon furin cleavage to allow EF and LF to bind (16,17). Several lines of evidence indicate that domain 2 (residues 259 -487) is involved in oligomerization and contains the loop that inserts into the membrane to form the channel through which the LF and EF enter the cytosol (16, 18 -20). Various amino acids in domain 3 (residues 488 -595) are necessary for oligomerization, and this has been the only function attributed to domain 3 to date (21,22). Domain 4 (residues 596 -735) is essential for binding to cellular receptor as indicated by several lines of...