Insights into the GTP/GDP cycle of RabX3, a novel GTPase from Entamoeba histolytica with tandem G-domains

Chandra, Mintu, Mukherjee, Madhumita, Srivastava, Vijay Kumar, Saito-Nakano, Yumiko, Nozaki, Tomoyoshi and Datta, Sunando (2014) Insights into the GTP/GDP cycle of RabX3, a novel GTPase from Entamoeba histolytica with tandem G-domains. Biochemistry, 53 7: 1191-1205. doi:10.1021/bi401428f


Author Chandra, Mintu
Mukherjee, Madhumita
Srivastava, Vijay Kumar
Saito-Nakano, Yumiko
Nozaki, Tomoyoshi
Datta, Sunando
Title Insights into the GTP/GDP cycle of RabX3, a novel GTPase from Entamoeba histolytica with tandem G-domains
Formatted title
Insights into the GTP/GDP cycle of RabX3, a novel GTPase from Entamoeba histolytica with tandem G-domains
Journal name Biochemistry   Check publisher's open access policy
ISSN 0006-2960
1520-4995
Publication date 2014-02-25
Sub-type Article (original research)
DOI 10.1021/bi401428f
Open Access Status Not yet assessed
Volume 53
Issue 7
Start page 1191
End page 1205
Total pages 15
Place of publication Washington, DC, United States
Publisher American Chemical Society
Language eng
Formatted abstract
Members of the small GTPase Ras superfamily regulate a host of systems through their ability to catalyze the GTP/GDP cycle. All family members reported thus far possess a single GTPase domain with a P-loop containing a nucleoside triphosphate hydrolase fold. Here for the first time we report a novel member from Entamoeba histolytica, EhRabX3, which harbors two GTPase domains in tandem and exhibits unique biochemical properties. A combination of biochemical and microcalorimetric studies revealed that EhRabX3 binds to a single guanine nucleotide through its N-terminal domain. Unlike most of the members of the Ras superfamily, the dissociation of the nucleotide from EhRabX3 is independent of Mg2+, perhaps indicating a novel mechanism of nucleotide exchange by this protein. We found that EhRabX3 is extremely sluggish in hydrolyzing GTP, and that could be attributed to its atypical nucleotide binding pocket. It harbors substitutions at two positions that confer oncogenicity to Ras because of impaired GTP hydrolysis. Engineering these residues into the conserved counterparts enhanced their GTPase activity by at least 20-fold. In contrast to most of the members of the Ras superfamily, EhRabX3 lacks the prenylation motif. Using indirect immunofluorescence and biochemical fractionation, we demonstrated that the protein is distributed all over the cytosol in amoebic trophozoites. Collectively, this unique ancient GTPase exhibits a striking evolutionary divergence from the other members of the superfamily.
Q-Index Code C1
Q-Index Status Provisional Code
Institutional Status Non-UQ

Document type: Journal Article
Sub-type: Article (original research)
Collection: Institute for Molecular Bioscience - Publications
 
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