Cloning, expression and characterization of glucokinase gene involved in the glucose-6-phosphate formation in <i>Staphylococcus aureus</i>

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Title	Cloning, expression and characterization of glucokinase gene involved in the glucose-6-phosphate formation in Staphylococcus aureus
Authors	Hanumanthu Prasanna Lakshmi¹, Sthanikam Yeswanth¹, Uppu Venkateswara Prasad¹, Dudipeta Vasu¹, Vimjam Swarupa¹, Pasupuleti Santhosh Kumar¹, Mangamoori Lakshmi Narasu² & Potukuchi Venkata Gurunadha Krishna Sarma¹*
Affiliation	¹Department of Biotechnology, Sri Venkateswara Institute of Medical Sciences, Tirupati, AP, India 517 507; ²Centre for Biotechnology, Institute of science and technology JNTU, Hyderabad, 500085
Email	sarmasvims@gmail.com; *Corresponding author
Article Type	Hypothesis
Date	Received January 23, 2013; Accepted January 28, 2013; Published February 21, 2013
Abstract	Glucose-6-phosphate (G-6-P) formation in Staphylococcus aureus is catalysed by glucokinase (glkA) gene under high glucose concentration leading to upregulation of various pathogenic factors; therefore the present study is aimed in the cloning and characterization of glk A gene from S. aureus ATCC12600. The glk A gene was cloned in the Sma I site of pQE 30, sequenced (Accession number: JN645812) and expressed in E. coli DH5a. The recombinant glk A expressed from the resultant glk A 1 clone was purified using nickel metal chelate chromatography, the pure enzyme gave single band in SDS-PAGE with molecular weight of 33kDa. The rglk A showed very high affinity to glucose Km 5.1±0.06mM with Hill coefficient of 1.66±0.032mM. Analysis of glucokinase sequence of S. aureus showed presence of typical ATP binding site and ROK motif CNCGRSGCIE. Sequentially and phylogenetically S. aureus glk A exhibited low identity with other bacterial glk A and 21% homology with human glucokinase (GCK). Functionally, S. aureus glk A showed higher rate of G-6-P formation compared to human GCK which may have profound role in the pathogenesis.
Keywords	glk A, pQE 30, ROK, Km.
Citation	*Lakshmi et al.* Bioinformation 9(4): 169-173 (2013)
Edited by	P Kangueane
ISSN	0973-2063
Publisher	Biomedical Informatics
License	This is an Open Access article which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. This is distributed under the terms of the Creative Commons Attribution License.