glpX gene of Mycobacterium tuberculosis: heterologous expression, purification, and enzymatic characterization of the encoded fructose 1,6-bisphosphatase II.

Abstract:

:The glpX gene (Rv1099c) of Mycobacterium tuberculosis (Mtb) encodes Fructose 1,6-bisphosphatase II (FBPase II; EC 3.1.3.11); a key gluconeogenic enzyme. Mtb possesses glpX homologue as the major known FBPase. This study explored the expression, purification and enzymatic characterization of functionally active FBPase II from Mtb. The glpX gene was cloned, expressed and purified using a two step purification strategy including affinity and size exclusion chromatography. The specific activity of Mtb FBPase II is 1.3 U/mg. The enzyme is oligomeric, followed Michaelis-Menten kinetics with an apparent km = 44 μM. Enzyme activity is dependent on bivalent metal ions and is inhibited by lithium and inorganic phosphate. The pH optimum and thermostability of the enzyme have been determined. The robust expression, purification and assay protocols ensure sufficient production of this protein for structural biology and screening of inhibitors against this enzyme.

authors

Gutka HJ,Rukseree K,Wheeler PR,Franzblau SG,Movahedzadeh F

doi

10.1007/s12010-011-9219-x

subject

Has Abstract

pub_date

2011-08-01 00:00:00

pages

1376-89

issue

8

eissn

0273-2289

issn

1559-0291

journal_volume

164

pub_type

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