Histidine kinases (HK) of bacterial two-component systems represent a hallmark of allosterism in proteins, being able to detect a signal through the sensor domain and transmit this information through the protein matrix to the kinase domain which, once active, autophosphorylates a specific histidine residue. Inactive-to-active transition results in a large conformational change that moves the kinase on top of the histidine. In the present work, we use several molecular simulation techniques (Molecular Dynamics, Hybrid QM/MM, and constant pH molecular dynamics) to study the activation and autophosphorylation reactions in L. plantarum WalK, a cis-acting HK. In agreement with previous results, we show that the chemical step requires tight coupling with the conformational step in order to maintain the histidine phosphoacceptor in the correct tautomeric state, with a reactive δ-nitrogen. During the conformational transition, the kinase domain is never released and walks along the HK helix axis, breaking and forming several conserved residue-based contacts. The phosphate transfer reaction is concerted in the transition state region and is catalyzed through the stabilization of the negative developing charge of transferring phosphate along the reaction.

Purpose Congenital hypopituitarism (CH) can present in isolation or with other birth defects. Mutations in multiple genes can cause CH, and the use of a genetic screening panel could establish the prevalence of mutations in known and candidate genes for this disorder. It could also increase the proportion of patients that receive a genetic diagnosis. Methods We conducted target panel genetic screening using single-molecule molecular inversion probes sequencing to assess the frequency of mutations in known hypopituitarism genes and new candidates in Argentina. We captured genomic DNA from 170 pediatric patients with CH, either alone or with other abnormalities. We performed promoter activation assays to test the functional effects of patient variants in LHX3 and LHX4. Results We found variants classified as pathogenic, likely pathogenic or with uncertain significance in 15.3% of cases. These variants were identified in known CH causative genes (LHX3, LHX4, GLI2, OTX2 and HESX1), in less frequently reported genes (FOXA2, BMP4, FGFR1, PROKR2, PNPLA6) and in new candidate genes (BMP2, HMGA2, HNF1A, NKX2-1). Conclusion In this work, we report the prevalence of mutations in known CH genes in Argentina and provide evidence for new candidate genes. We show that CH is a genetically heterogeneous disease with high phenotypic variation and incomplete penetrance, and our results support the need for further gene discovery for CH. Identifying population-specific pathogenic variants will improve the capacity of genetic data to predict eventual clinical outcomes.