A conserved P-loop anchor limits the structural dynamics that mediate nucleotide dissociation in EF-Tu

Abstract

The phosphate-binding loop (P-loop) is a conserved sequence motif found in mononucleotide-binding proteins. Little is known about the structural dynamics of this region and its contribution to the observed nucleotide binding properties. Understanding the underlying design principles is of great interest for biomolecular engineering applications. We have used rapid-kinetics measurements in vitro and molecular dynamics (MD) simulations in silico to investigate the relationship between GTP-binding properties and P-loop structural dynamics in the universally conserved Elongation Factor (EF) Tu. Analysis of wild type EF-Tu and variants with substitutions at positions in or adjacent to the P-loop revealed a correlation between P-loop flexibility and the entropy of activation for GTP dissociation. The same variants demonstrate more backbone flexibility in two N-terminal amino acids of the P-loop during force-induced EF-Tu·GTP dissociation in Steered Molecular Dynamics simulations. Amino acids Gly18 and His19 are involved in stabilizing the P-loop backbone via interactions with the adjacent helix C. We propose that these P-loop/helix C interactions function as a conserved P-loop anchoring module and identify the presence of P-loop anchors within several GTPases and ATPases suggesting their evolutionary conservation.

Glycyrrhetinic acid binds to the conserved P-loop region and interferes with the interaction of RAS-effector proteins

“…Conformational changes in the P-loop may be a common strategy contributing to GEF-stimulated nucleotide exchange in G-proteins. Thus, Structural disruption of the P-loop is likely the essential mechanism underlying the sharply reduced catalytic function of GEFs for GDP release and GDP–GTP exchange 53 , which leads to changes in affinity and kinetics of interaction with effectors.…”

Conformational Destabilization of Immunoglobulin G Increases the Low pH Binding Affinity with the Neonatal Fc Receptor

“…A possible reason relates to the unknown contribution(s) of conformational dynamics (1). Although not repre-sented in static crystal structures, conformational dynamics have been shown to modulate dissociation rates (2)(3)(4)(5), influence substrate transport (6), and even regulate protein activity (7,8).…”

Loss of phosphatase activity in PTEN (phosphatase and tensin homolog deleted on chromosome ten) results in endometrial carcinoma in humans: An in-silico study

“…The function of the amino acids within P-loop allow for anchoring and help to detect the P-loop anchors in the several GTPase and ATPase. Glycine and histidine stabilize the major structure of P-loop by creating the interaction between adjacent helix C (Mercier et al, 2015).…”

“…There are many studies, which showed the repression of PTEN gene expression, rather than only mutations and deletions, are associated with tumorigenesis. Studies showed, besides mutations in some important domains within PTEN, various molecular mechanisms, which can cause changes in PTEN protein levels, location, and/or enzymatic activity can have negative effects on PTEN normal functioning in sporadic cancers, and other diseases (Stambolic and Woodgett, 2006;Milella et al, 2015). Decreased PTEN transcript levels were found to be associated with PTEN promoter methylation in melanoma.…”