Characterization of the universally conserved ATPase YchF using an in vitro and in silico approach
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Date
2014
Authors
Rosler, Kirsten S
University of Lethbridge. Faculty of Arts and Science
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Publisher
Lethbridge, Alta : University of Lethbridge, Dept. of Chemistry and Biochemistry
Abstract
GTPases perform a myriad of functions within the cell, from protein
synthesis to cellular signaling. Of all known GTPases, only eight are conserved
across all three domains of life. YchF is one of the eight universally conserved
GTPases, however its cellular function is poorly understood. YchF differs from the
classical GTPases in that it has a higher affinity for ATP than for GTP and
functions as an ATPase. As a HAS-GTPase, YchF does not possess the
canonical catalytic glutamine required for nucleotide hydrolysis, urging the
question of how does YchF hydrolyze ATP. Here we have used molecular
dynamics simulations (in silico) and biochemical experiments (in vitro) to identify
an amino acid, histidine 114, essential for ATP hydrolysis in YchF. His 114 is
located in a flexible loop of the G-domain of YchF, which shows nucleotidedependent
conformations in silico. The findings reported indicate that the 70S
ribosome can stimulate the ATPase activity of YchF by directly participating in
catalysis as well as helping to position the catalytic histidine residue. Additionally,
nucleotide binding and dissociation rate constants have been determined in the
presence and absence of Mg2+ in order to further understand the functional cycle
of YchF.
Description
Keywords
YchF , HAS-GTPase , ATPase , catalytic mechanism , nucleotide binding