Investigation of a Putative Arginine Kinase from Desulfobacterium autotrophicum: Evolutionary Implications for the Bacterial Arginine Kinases

Christopher DeMoll, The College of Wooster

Abstract

A putative arginine kinase from Desulfobacterium autotrophicum was cloned into a pET45 vector equipped with a Histag. The Dauto-AK was then expressed in E. coli and purified using Ni2+ affinity-chromatography. HPLC was used to confirm synthesis of phosphoarginine from the reaction of the AK with the two substrates MgATP and arginine. Attempts to use ITC to characterize the Dauto-AK protein were unsuccessful. In addition, an MSA and phylogenetic tree were constructed to include all eight members of the bacterial AKs. Non-conserved residues were discovered in the Region-95 area of the AKs from Desulfobacterium, Ahrensia, and Myxococcus that usually exhibits full conservation amongst PKs, as it is thought to play a role in guanidino substrate specificity. The genomes of both Myxococcus and Desulfobacterium were discovered to have an ABC-type glycine betaine/proline transport system, ATP-binding protein in the immediate vicinity of their AK coding sequences. However, these proteins did not exhibit enough relation to suggest that the AKs came from a similar horizontal gene transfer event. All evidence obtained here corroborates the pre-existing theory that each instance of a bacterial AK is the result of an independent horizontal gene transfer event. Preliminary STRING searches revealed possible proteins for the investigation of protein-protein interactions in bacterial AKs. Both an arginyl-tRNA synthetase, and an argininosuccinate lyase were identified as having potential interactions with the bacterial AK from Sulfurovum.

 

© Copyright 2011 Christopher DeMoll