PknB-Mediated Phosphorylation of a Novel Substrate, N-Acetylglucosamine-1-Phosphate Uridyltransferase, Modulates Its Acetyltransferase Activity

Amit Parikh, Sunil Kumar Verma, Shazia Khan, Balaji Prakash, Vinay Kumar Nandicoori

Research output: Contribution to journalArticle

79 Citations (Scopus)

Abstract

Identifying direct targets of kinases and determining how their activities are regulated are central to understanding how they generate biological responses. Genetic and biochemical studies have shown that Mycobacterium tuberculosis serine/threonine protein kinases PknA and PknB play a role in modulating cell shape and possibly cell division. In this report, we show that the enzyme N-acetylglucosamine-1-phosphate uridyltransferase (GlmU) of M. tuberculosis is a novel substrate of PknB and is phosphorylated on threonine residues. GlmU carries out two important biochemical activities: a C-terminal domain catalyzes the transfer of acetyl group from acetyl coenzyme A to glucosamine-1-phosphate to produce N-acetylglucosamine-1-phosphate, which is converted into UDP-N-acetylglucosamine by the transfer of uridine 5'-monophosphate (from uridine 5′-triphosphate), a reaction catalyzed by the N-terminal domain. We determined the crystal structures of GlmU in apo form and UDP-N-acetylglucosamine-bound form, and analyzed them to identify threonine residues that may be accessible to PknB. The structure shows a two-domain architecture, with an N-terminal domain having an α/β-like fold and with a C-terminal domain that forms a left-handed parallel β-helix structure. Kinase assays with PknB using the N- and C-terminal domains of GlmU as substrates illustrated that PknB phosphorylates GlmU in the C-terminal domain. Furthermore, mutational studies reveal one of the five threonines present in region 414-439 to be phosphorylated by PknB. Structural and biochemical analyses have shown the significance of a variable C-terminal tail in regulating acetyltransferase activity. Notably, we demonstrate that although PknB-mediated phosphorylation of GlmU does not affect its uridyltransferase activity, it significantly modulates the acetyltransferase activity. These findings imply a role for PknB in regulating peptidoglycan synthesis by modulating the acetyltransferase activity of GlmU.

Original languageEnglish (US)
Pages (from-to)451-464
Number of pages14
JournalJournal of Molecular Biology
Volume386
Issue number2
DOIs
StatePublished - Feb 20 2009
Externally publishedYes

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Acetyltransferases
Threonine
Uridine Diphosphate N-Acetylglucosamine
Phosphorylation
Mycobacterium tuberculosis
Phosphotransferases
Uridine Monophosphate
Uridine Triphosphate
Acetyl Coenzyme A
Peptidoglycan
Cell Shape
Protein-Serine-Threonine Kinases
Cell Division
Molecular Biology
Enzymes
UDPacetylglucosamine pyrophosphorylase

Keywords

  • GlmU
  • Mycobacterium tuberculosis
  • peptidoglycan
  • phosphorylation
  • PknB

ASJC Scopus subject areas

  • Molecular Biology

Cite this

PknB-Mediated Phosphorylation of a Novel Substrate, N-Acetylglucosamine-1-Phosphate Uridyltransferase, Modulates Its Acetyltransferase Activity. / Parikh, Amit; Verma, Sunil Kumar; Khan, Shazia; Prakash, Balaji; Nandicoori, Vinay Kumar.

In: Journal of Molecular Biology, Vol. 386, No. 2, 20.02.2009, p. 451-464.

Research output: Contribution to journalArticle

Parikh, Amit ; Verma, Sunil Kumar ; Khan, Shazia ; Prakash, Balaji ; Nandicoori, Vinay Kumar. / PknB-Mediated Phosphorylation of a Novel Substrate, N-Acetylglucosamine-1-Phosphate Uridyltransferase, Modulates Its Acetyltransferase Activity. In: Journal of Molecular Biology. 2009 ; Vol. 386, No. 2. pp. 451-464.
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