Distinct GDP/GTP bound states of the tandem G-domains of EngA regulate ribosome binding

EngA, a unique GTPase containing a KH-domain preceded by two consecutive G-domains, displays distinct nucleotide binding and hydrolysis activities. So far, Escherichia coli EngA is reported to bind the 50S ribosomal subunit in the guanosine-5′-trihosphate (GTP) bound state. Here, for the first time, using mutations that allow isolating the activities of the two G-domains, GD1 and GD2, we show that apart from 50S, EngA also binds the 30S and 70S subunits. We identify that the key requirement for any EngA–ribosome association is GTP binding to GD2. In this state, EngA displays a weak 50S association, which is further stabilized when GD1 too binds GTP. Exchanging bound GTP with guanosine-5′-diphosphate (GDP), at GD1, results in interactions with 50S, 30S and 70S. Therefore, it appears that GD1 employs GTP hydrolysis as a means to regulate the differential specificity of EngA to either 50S alone or to 50S, 30S and 70S subunits. Furthermore, using constructs lacking either GD1 or both GD1 and GD2, we infer that GD1, when bound to GTP and GDP, adopts distinct conformations to mask or unmask the 30S binding site on EngA. Our results suggest a model where distinct nucleotide-bound states of the two G-domains regulate formation of specific EngA–ribosome complexes

Role of GTPases in ribosome assembly

GTPases are a universally conserved class of regulatory proteins involved in such diverse cellular functions as signal transduction, translation, cytoskeleton formation, and intracellular transport. GTPases are also required for ribosome assembly in eukaryotes and bacteria, where they present themselves as possible regulatory molecules. Strikingly, in bacteria they represent the largest class of essential assembly factors. A review of their common structural, biochemical and genetic interactions is presented and integrated with models for their function in ribosome assembly. © 2007 Wiley Periodicals, Inc. Biopolymers 87: 1–11, 2007 This article was originally published online as an accepted preprint. The “Published Online” date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at [email protected] Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/56108/1/20762_ftp.pd

Deciphering the Catalytic Machinery in 30S Ribosome Assembly GTPase YqeH

YqeH, a circularly permuted GTPase (cpGTPase), which is conserved across bacteria and eukaryotes including humans is important for the maturation of small (30S) ribosomal subunit in Bacillus subtilis. Recently, we have shown that it binds 30S in a GTP/GDP dependent fashion. However, the catalytic machinery employed to hydrolyze GTP is not recognized for any of the cpGTPases, including YqeH. This is because they possess a hydrophobic substitution in place of a catalytic glutamine (present in Ras-like GTPases). Such GTPases were categorized as HAS-GTPases and were proposed to follow a catalytic mechanism, different from the Ras-like proteins.MnmE, another HAS-GTPase, but not circularly permuted, utilizes a potassium ion and water mediated interactions to drive GTP hydrolysis. Though the G-domain of MnmE and YqeH share only approximately 25% sequence identity, the conservation of characteristic sequence motifs between them prompted us to probe GTP hydrolysis machinery in YqeH, by employing homology modeling in conjunction with biochemical experiments. Here, we show that YqeH too, uses a potassium ion to drive GTP hydrolysis and stabilize the transition state. However, unlike MnmE, it does not dimerize in the transition state, suggesting alternative ways to stabilize switches I and II. Furthermore, we identify a potential catalytic residue in Asp-57, whose recognition, in the absence of structural information, was non-trivial due to the circular permutation in YqeH. Interestingly, when compared with MnmE, helix alpha2 that presents Asp-57 is relocated towards the N-terminus in YqeH. An analysis of the YqeH homology model, suggests that despite such relocation, Asp-57 may facilitate water mediated catalysis, similarly as the catalytic Glu-282 of MnmE. Indeed, an abolished catalysis by D57I mutant supports this inference.An uncommon means to achieve GTP hydrolysis utilizing a K(+) ion has so far been demonstrated only for MnmE. Here, we show that YqeH also utilizes a similar mechanism. While the catalytic machinery is similar in both, mechanistic differences may arise based on the way they are deployed. It appears that K(+) driven mechanism emerges as an alternative theme to stabilize the transition state and hydrolyze GTP in a subset of GTPases, such as the HAS-GTPases

The chloroplast protein BPG2 functions in brassinosteroid-mediated post-transcriptional accumulation of chloroplast rRNA

Brassinazole (Brz) is a specific inhibitor of the biosynthesis of brassinosteroids (BRs), which regulate plant organ and chloroplast development. We identified a recessive pale green Arabidopsis mutant, bpg2-1 (Brz-insensitive-pale green 2-1) that showed reduced sensitivity to chlorophyll accumulation promoted by Brz in the light. BPG2 encodes a chloroplast-localized protein with a zinc finger motif and four GTP-binding domains that are necessary for normal chloroplast biogenesis. BPG2-homologous genes are evolutionally conserved in plants, green algae and bacteria. Expression of BPG2 is induced by light and Brz. Chloroplasts of the bpg2-1 mutant have a decreased number of stacked grana thylakoids. In bpg2-1 and bpg2-2 mutants, there was no reduction in expression of rbcL and psbA, but there was abnormal accumulation of precursors of chloroplast 16S and 23S rRNA. Chloroplast protein accumulation induced by Brz was suppressed by the bpg2 mutation. These results indicate that BPG2 plays an important role in post-transcriptional and translational regulation in the chloroplast, and is a component of BR signaling.publishersversionPeer reviewe

A review of friction models in interacting joints for durability design.

This paper presents a comprehensive review of friction modelling to provide an understanding of design for durability within interacting systems. Friction is a complex phenomenon and occurs at the interface of two components in relative motion. Over the last several decades, the effects of friction and its modelling techniques have been of significant interests in terms of industrial applications. There is however a need to develop a unified mathematical model for friction to inform design for durability within the context of varying operational conditions. Classical dynamic mechanisms model for the design of control systems has not incorporated friction phenomena due to non-linearity behaviour. Therefore, the tribological performance concurrently with the joint dynamics of a manipulator joint applied in hazardous environments needs to be fully analysed. Previously the dynamics and impact models used in mechanical joints with clearance have also been examined. The inclusion of reliability and durability during the design phase is very important for manipulators which are deployed in harsh environmental and operational conditions. The revolute joint is susceptible to failures such as in heavy manipulators these revolute joints can be represented by lubricated conformal sliding surfaces. The presence of pollutants such as debris and corrosive constituents has the potential to alter the contacting surfaces, would in turn affect the performance of revolute joints, and puts both reliability and durability of the systems at greater risks of failure. Key literature is identified and a review on the latest developments of the science of friction modelling is presented here. This review is based on a large volume of knowledge. Gaps in the relevant field have been identified to capitalise on for future developments. Therefore, this review will bring significant benefits to researchers, academics and industrial professionals

A comprehensive survey of the analytical, numerical and experimental methodologies for dynamics of multibody mechanical systems with clearance or imperfect joints

"Available online 19 December 2017"A comprehensive survey of the literature of the most relevant analytical, numerical, and experimental approaches for the kinematic and dynamic analyses of multibody mechanical systems with clearance joints is presented in this review. Both dry and lubricated clearance joints are addressed here, and an effort is made to include a large number of research works in this particular field, which have been published since the 1960′s. First, the most frequently utilized methods for modeling planar and spatial multibody mechanical systems with clearance joints are analyzed, and compared. Other important phenomena commonly associated with clearance joint models, such as wear, non-smooth behavior, optimization and control, chaos, and uncertainty and links’ flexibility, are then discussed. The main assumptions procedures and conclusions for the different methodologies are also examined and compared. Finally, future developments and new applications of clearance joint modeling and analysis are highlighted.This research was supported in part by the China 111 Project (B16003) and the National Natural Science Foundation of China under Grants 11290151, 11472042 and 11221202. The work was also supported by the Portuguese Foundation for Science and Technology with the reference project UID/EEA/04436/2013, by FEDER funds through the COMPETE 2020 – Programa Operacional Competitividade e Internacionalização (POCI) with the reference project POCI-01-0145-FEDER-006941.info:eu-repo/semantics/publishedVersio

Ein Ssitrag zur Kenntnis der chronischen peripheren Verduennung und Ektasie der Hornhaut.

Munch en. 190.OPLADEN-RUG0