Bile salt hydrolase (BSH) is an enzyme produced by the intestinal microflora that catalyzes the deconjugation of glycine-or taurine-linked bile salts. The crystal structure of BSH reported here from Bifidobacterium longum reveals that it is a member of N-terminal nucleophil hydrolase structural superfamily possessing the characteristic ␣␤␤␣ tetra-lamellar tertiary structure arrangement. Site-directed mutagenesis of the catalytic nucleophil residue, however, shows that it has no role in zymogen processing into its corresponding active form. Substrate specificity was studied using Michaelis-Menten and inhibition kinetics and fluorescence spectroscopy. These data were compared with the specificity profile of BSH from Clostridium perfrigens and pencillin V acylase from Bacillus sphaericus, for both of which the three-dimensional structures are available. Comparative analysis shows a gradation in activity toward common substrates, throwing light on a possible common route toward the evolution of pencillin V acylase and BSH.

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Co3O4, which crystallizes in the spinel phase at temperatures much higher than ambient temperatures, orders antiferromagnetically below its Neel temperature (∼40 K). However, in nanosize, it shows ferromagnetic ordering due to surface canting, disorder, imperfect oxygen atom coordination, etc. Here, for the first time, we report the synthesis of single-crystalline, ferromagnetic Co3O4 nanoparticles functionalized with proteins (size 5–7 nm) using cobalt acetate as precursor at room temperature in aqueous medium by a metal-tolerant marine bacterium isolated from the coast of the Arabian Sea. Energetically unfavorable change in the oxidation state of Co atoms from (+2 to +3) during synthesis by the bacterial strain was evidenced by X-ray photoelectron spectroscopy. The effect on the morphology of bacterial cells after exposure to the cobalt acetate was imaged by scanning electron microscopy showing cooperative, self-organized, structured colony formation, possibly due to the interbacterial communication under external stress.

Virulence pathways in gram-negative pathogenic bacteria are regulated by quorum sensing mechanisms, through the production and sensing of N-acylhomoserine lactone (AHL) signal molecules. Enzymatic degradation of AHLs leading to attenuation of virulence (quorum quenching) could pave the way for the development of new antibacterials. Penicillin V acylases (PVAs) belong to the Ntn hydrolase superfamily, together with AHL acylases. PVAs are exploited widely in the pharmaceutical industry, but their role in the natural physiology of their native microbes is not clearly understood. This report details the characterization of AHL degradation activity by homotetrameric PVAs from two gram-negative plant pathogenic bacteria, Pectobacterium atrosepticum (PaPVA) and Agrobacterium tumefaciens (AtPVA). Both the PVAs exhibited substrate specificity for degrading long-chain AHLs. Exogenous addition of these enzymes into Pseudomonas aeruginosa greatly diminished the production of elastase and pyocyanin and biofilm formation and increased the survival rate in an insect model of acute infection. Subtle structural differences in the PVA active site that regulate specificity for acyl chain length have been characterized, which could reflect the evolution of AHL-degrading acylases in relation to the environment of the bacteria that produce them and also provide strategies for enzyme engineering. The potential for using these enzymes as therapeutic agents in clinical applications and a few ideas about their possible significance in microbial physiology have also been discussed.

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The formation of biocomposite films of the pharmaceutically important enzyme penicillin G acylase (PGA) and fatty lipids under enzyme-friendly conditions is described. The approach involves a simple beaker-based diffusion protocol wherein the enzyme diffuses into the lipid film during immersion in the enzyme solution, thereby leading to the formation of a biocomposite film. The incorporation of the enzyme in both cationic as well as anionic lipids suggests the important role of secondary interactions such as hydrophobic and hydrogen bonding in the enzyme immobilization process. The kinetics of formation of the enzyme-lipid biocomposites has been studied by quartz crystal microgravimentry (QCM) measurements. The stability of the enzyme in the lipid matrix was confirmed by Fourier transform infrared spectroscopy (FTIR) and biocatalytic activity measurements. Whereas the biological activity of the lipid-immobilized enzyme was marginally higher than that of the free enzyme, the biocomposite film exhibited increased thermal/temporal stability. Particularly exciting was the observation that the biocomposite films could be reused in biocatalysis reactions without significant loss in activity, which indicates potentially exciting biomedical/industrial application of these films.

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