Adenosine-5'-carboxylic acid peptidyl derivatives as inhibitors of protein kinases.

A new class of protein kinase bisubstrate-analog inhibitors was designed on the basis of adenosine-5'-carboxylic acid derivatives, where a short peptide was attached to the 5'-carbon atom of the adenosine sugar moiety via a linker chain. The potency and selectivity of these inhibitors were adjusted by relevant combination of these structural fragments, resembling the structure of the bisubstrate complex of the peptide phosphorylation reaction.     

Controls for Phylogeny and Robust Analysis in Pareto Task Inference

Understanding the tradeoffs faced by organisms is a major goal of evolutionary biology. One of the main approaches for identifying these tradeoffs is Pareto task inference (ParTI). Two recent papers claim that results obtained in ParTI studies are spurious due to phylogenetic dependence (Mikami T, Iwasaki W. 2021. The flipping t-ratio test: phylogenetically informed assessment of the Pareto theory for phenotypic evolution. Methods Ecol Evol. 12(4):696–706) or hypothetical p-hacking and population-structure concerns (Sun M, Zhang J. 2021. Rampant false detection of adaptive phenotypic optimization by ParTI-based Pareto front inference. Mol Biol Evol. 38(4):1653–1664). Here, we show that these claims are baseless. We present a new method to control for phylogenetic dependence, called SibSwap, and show that published ParTI inference is robust to phylogenetic dependence. We show how researchers avoided p-hacking by testing for the robustness of preprocessing choices. We also provide new methods to control for population structure and detail the experimental tests of ParTI in systems ranging from ammonites to cancer gene expression. The methods presented here may help to improve future ParTI studies.     

Determination of the physiological dimer interface of the PhoQ sensor domain

PhoQ is the transmembrane sensor kinase of the phoPQ two-component system, which detects and responds to divalent cations and antimicrobial peptides and can trigger bacterial virulence. Despite their ubiquity and importance in bacterial signaling, the structure and molecular mechanism of the sensor kinases is not fully understood. Frequently, signals are transmitted from a periplasmic domain in these proteins to the cytoplasmic kinase domains via an extended dimeric interface, and the PhoQ protein would appear to follow this paradigm. However, the isolated truncated periplasmic domain of PhoQ dimerizes poorly, so it has been difficult to distinguish the relevant interface in crystal structures of the PhoQ periplasmic domain. Thus, to determine the arrangement of the periplasmic domains of Escherichia coli PhoQ in the physiological homodimer, disulfide-scanning mutagenesis was used. Single cysteine substitutions were introduced along the N-terminal helix of the periplasmic region, and the degree of cross-linking in each protein variant was determined by Western blotting and immunodetection. The results were subjected to periodicity analysis to generate a profile that provides information concerning the C^β distances between corresponding residues at the interface. This profile, together with a rigid-body search procedure, side-chain placement, and energy minimization, was used to build a model of the dimer arrangement. The final model proved to be highly compatible with one of the PhoQ crystal structures, 3BQ8, indicating that 3BQ8 is representative of the physiological arrangement. The model of the periplasmic region is also compatible with a full-length PhoQ protein in which a four-helix bundle forms in the membrane. The membrane four-helix bundle has been proposed for other sensor kinases and is thought to have a role in the mechanism of signal transduction; our model supports the idea that signaling through a membrane four-helix bundle is a widespread mechanism in the transmembrane sensor kinases.     

Synthesis and evaluation of novel dipeptidyl benzoyloxymethyl ketones as caspase inhibitors

We describe novel peptide-based caspase inhibitors. Potent and comparatively selective compounds containing a dipeptide scaffold and a substituted oxymethyl ketone as a warhead were developed. The newly synthesized compounds were tested for inhibition in in vitro enzymatic assays of caspases-1, -3, -6, -8, and -9. The benzyloxycarbonyl-phenylglycyl-aspartyl benzoyloxymethyl ketone (Z-Phg-Asp-CH2OCO-Ph, coded as HU44) was the most potent inhibitor of caspase-1 and caspase-3. Of several analogs of HU44 that were made, the beta-Asp methyl ester (2) is an effective inhibitor against caspase-3 and caspase-8, and less effective against caspase-1. These compounds did not inhibit caspase-6 and caspase-9 significantly.