Staphylococcal PknB as the First Prokaryotic Representative of the Proline-Directed Kinases

Abstract

In eukaryotic cell types, virtually all cellular processes are under control of proline-directed kinases and especially MAP kinases. Serine/threonine kinases in general were originally considered as a eukaryote-specific enzyme family. However, recent studies have revealed that orthologues of eukaryotic serine/threonine kinases exist in bacteria. Moreover, various pathogenic species, such as Yersinia and Mycobacterium, require serine/threonine kinases for successful invasion of human host cells. The substrates targeted by bacterial serine/threonine kinases have remained largely unknown. Here we report that the serine/threonine kinase PknB from the important pathogen Staphylococcus aureus is released into the external milieu, which opens up the possibility that PknB does not only phosphorylate bacterial proteins but also proteins of the human host. To identify possible human targets of purified PknB, we studied in vitro phosphorylation of peptide microarrays and detected 68 possible human targets for phosphorylation. These results show that PknB is a proline-directed kinase with MAP kinase-like enzymatic activity. As the potential cellular targets for PknB are involved in apoptosis, immune responses, transport, and metabolism, PknB secretion may help the bacterium to evade intracellular killing and facilitate its growth. In apparent agreement with this notion, phosphorylation of the host-cell response coordinating transcription factor ATF-2 by PknB was confirmed by mass spectrometry. Taken together, our results identify PknB as the first prokaryotic representative of the proline-directed kinase/MAP kinase family of enzymes.

Phosphorylation controls the functioning ofStaphylococcus aureusisocitrate dehydrogenase – favours biofilm formation

“…This plays crucial role in the regulation of various metabolic processes in the organism as well as expression of various virulence factors and by passing the host immune system, thus helping the pathogen in colonization in any anatomical locales in human host. This condition is further aided by the presence of secretary PknB and alkaline phosphatase [21][22][23] . In S. aureus cell wall, exopolysaccharide and PIA molecule biosynthesis are primarily regulated by PknB.…”

“…In the present study we have cloned, sequenced, expressed and characterized PknB and alkaline phosphatase III (SAOV 2675) genes of S. aureus ATCC12600. The PknB sequence showed well-conserved PASTA domains which sense environmental signals by triggering eukaryote-like N-terminal catalytic domain to autophosphorylate and hence activate different downstream signalling cascades which help to regulate physiology of the organism 11,[15][16][17]21,34,37 . The results of the current study also explain unambiguously how IDH become completely inactive on phosphorylation with PknB and are restored upon dephosphorylation with alp III (Table 4).…”

“…This pathogen survives in the human host mostly as biofilms contributing resistance to several antibiotics making S. aureus as a multidrug resistant strain 11,[15][16][17][18][19][20] . Further, in this organism, PknB is actively present in the external secretions, and along with phosphatases they interact with vast range of host tissue proteins aiding this pathogen to colonize in any anatomical locales in human host 21,22 . In S. aureus, the dephosphorylation is carried out by phosphatases which are present along with phosphokinases.…”

Membrane interactions and self‐association of components of the Ess/Type VII secretion system of Staphylococcus aureus

“…SDS PAGE was carried out using Bis‐Tris gels as described previously 3, and western blotting was performed following standard protocols with the following antibody dilutions: α‐EsxA 3 1 : 2500, α‐EsxC 3 1 : 2000, α‐EsaA 3 1 : 10 000, α‐EssB 3 1 : 10 000, α‐EssC 3 1 : 10 000 and α‐TrxA 13 1 : 10 000.…”

Systems medicine approaches for peptide array-based protein kinase profiling: progress and prospects

“…An analysis of the Physarum polycephalum genome demonstrated extensive use of the twocomponent-type prokaryotic kinases as well of metazoantype tyrosine kinase-like genes [17]. Approaches involving peptide arrays for characterizing the enzymatic activity towards substrates are now starting to be used for characterizing these elements [18] and are used to study modulation of their activity during infection [19]. As antibiotic resistance is emerging as serious health problem requiring radically novel approaches (e.g.…”