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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 5.10.06mM with Hill coefficient of 1.660.032mM. 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.