The review highlights the recent progress and challenges in developing a family of nitrile reductases as biocatalysts for nitrile-to-amine transformation.

c Type IV pili (T4P) are polar surface structures that play important roles in bacterial motility, biofilm formation, and pathogenicity. The protein FimX and its orthologs are known to mediate T4P formation in the human pathogen Pseudomonas aeruginosa and some other bacterial species. It was reported recently that FimX XAC2398 from Xanthomonas axonopodis pv. citri interacts with PilZ XAC1133 directly through the nonenzymatic EAL domain of FimX XAC2398 . Here we present experimental data to reveal that the strong interaction between FimX XAC2398 and PilZ XAC1133 is not conserved in P. aeruginosa and likely other Pseudomonas species. In vitro and in vivo binding experiments showed that the interaction between FimX and PilZ in P. aeruginosa is below the measurable limit. Surface plasmon resonance assays further confirmed that the interaction between the P. aeruginosa proteins is at least more than 3 orders of magnitude weaker than that between the X. axonopodis pv. citri pair. The N-terminal lobe region of FimX XAC2398 was identified as the binding surface for PilZ XAC1133 by amide hydrogen-deuterium exchange and site-directed mutagenesis studies. Lack of several key residues in the N-terminal lobe region of the EAL domain of FimX is likely to account for the greatly reduced binding affinity between FimX and PilZ in P. aeruginosa. All together, the results suggest that the interaction between PilZ and FimX in Xanthomonas species is not conserved in P. aeruginosa due to the evolutionary divergence among the FimX orthologs. The precise roles of FimX and PilZ in bacterial motility and T4P biogenesis are likely to vary among bacterial species.

Messenger bagged: The design of a fluorophore-labeled protein biosensor for the bacterial messenger cyclic di-GMP is described. The biosensor responds to c-di-GMP with sub-micromolar sensitivity in a real-time fashion. The biosensor can be used for enzyme assays for diguanylate cyclases and c-di-GMP phosphodiesterases as well as the high-throughput screening of inhibitors.

The cover picture shows the binding of the bacterial messenger cyclic di-GMP (c-di-GMP) to a protein-based fluorescent biosensor. C-di-GMP plays important roles in many pathogenic bacteria by mediating biofilm formation and virulence (a biofilm formed by P. aeruginosa is shown in the background), and bacterial cells usually contain multiple diguanylate cyclases (DGCs) and phosphodiesterases (PDEs) to regulate the cellular concentration of c-di-GMP. Certain proteins are now being considered as novel targets for the development of antibacterial agents. The development of facile methods of assaying enzymatic activity and screening inhibitors will facilitate the lead discovery processes. On p. 2753 ff, Z.-X. Liang et al. describe the design of a fluorescent biosensor that responds to c-di-GMP with sub-micromolar sensitivity in a real-time fashion. The biosensor can be used in enzyme assays for DGCs and c-di-GMP PDEs as well as the highthroughput screening of inhibitors.