A Serine/Threonine Protein Kinase fromMycobacterium tuberculosis

Abstract

Genomic DNA sequencing in the vicinity of the pstA-1 gene from Mycobacterium tuberculosis allowed us to clone, sequence and identify a gene encoding a 70-kDa protein. The size of the protein was confirmed by in vitru coupled transcriptiodtranslation. Its N-terminal domain shows extensive sequence similarity with the catalytic domain of eukaryotic serinelthreonine protein kinases, and the protein was therefore called Mbk (mycobacterial protein kinase). The deduced amino acid sequence contains two transmembrane segments, which flank a highly repetitive region, suggesting a receptor-like anchoring. The mbk gene was overexpressed in Escherichia coli and the gene product (Mbk) was purified as a fusion protein with gluthatione S-transferase. Recombinant Mbk was found to be autophosphorylated on threonine residues and capable of phosphorylating myelin basic proteins from bovine brain and histones from calf thymus on serine residues, both in a manganese-dependent manner. The phosphorylation of myelin basic proteins by Mbk was inhibited by calcium and by staurosporine, a widely used inhibitor of eukaryotic protein serinekhreonine kinases. A similar gene was found in Mycobacterium bovis BCG DNA by Southern blot analysis. Its expression was detected in cultures of M. bovis BCG by reverse transcriptasel PCR. Although its biological role is unknown, it is the first serine/threonine protein kinase characterized in Mycobacteria.

The Mycobacterium tuberculosis β-Ketoacyl-Acyl Carrier Protein Synthase III Activity Is Inhibited by Phosphorylation on a Single Threonine Residue

“…PknB, PknK, and PknL did not phosphorylate mtFabH, which may be due, at least for PknK and PknL, to their very low autokinase activity. STPKs usually migrate as diffuse bands, reflecting the different levels of phosphorylation for each isoform, and this aberrant profile of migration of M. tuberculosis kinases has already been reported in earlier studies (17,(37)(38)(39). These results are consistent with the fact that KasA and KasB were also found to be efficiently phosphorylated by PknA and PknF (24).…”

From the Characterization of the Four Serine/Threonine Protein Kinases (PknA/B/G/L) of Corynebacterium glutamicum toward the Role of PknA and PknB in Cell Division

“…All these bands were analyzed by mass spectrometry, which confirmed the identity of the GST chimeric proteins (data not shown). An aberrant migration property has been observed previously for M. tuberculosis kinases, such as PknD, PknE, PknL, and PknH (20,31,39,40), as well as for Pkn9 and Pkn6, two Myxococcus xanthus STPKs (41,42). It was demonstrated that this specific pattern of migration was because of the different phosphorylated isoforms.…”

The Mycobacterium tuberculosis serine/threonine kinases PknA and PknB: substrate identification and regulation of cell shape

“…The M. tuberculosis genome contains genes encoding 11 STPKs, all of which are in the PKN2 family of prokaryotic protein kinases that are most closely related to the eukaryotic STPKs (Cole et al 1998;Leonard et al 1998). To date, these kinases have been characterized to a limited extent, and no specific substrates have been identified (Peirs et al 1997;Av-Gay et al 1999; Av-Gay and Everett 2000; Koul et al 2001;Chaba et al 2002).…”