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Scientific article
English

Intersubunit cross-talk in pyridoxal 5'-phosphate synthase, coordinated by the C terminus of the synthase subunit

Published inThe Journal of biological chemistry, vol. 284, no. 12, p. 7706-7718
Publication date2009
Abstract

Vitamin B(6) is essential in all organisms, due to its requirement as a cofactor in the form of pyridoxal 5'-phosphate (PLP) for key metabolic enzymes. It can be synthesized de novo by either of two pathways known as deoxyxylulose 5-phosphate (DXP)-dependent and DXP-independent. The DXP-independent pathway is the predominant pathway and is found in most microorganisms and plants. A glutamine amidotransferase consisting of the synthase Pdx1 and its glutaminase partner, Pdx2, form a complex that directly synthesizes PLP from ribose 5-phosphate, glyceraldehyde 3-phosphate, and glutamine. The protein complex displays an ornate architecture consisting of 24 subunits, two hexameric rings of 12 Pdx1 subunits to which 12 Pdx2 subunits attach, with the glutaminase and synthase active sites remote from each other. The multiple catalytic ability of Pdx1, the remote glutaminase and synthase active sites, and the elaborate structure suggest regulation of activity on several levels. A missing piece in deciphering this intricate puzzle has been information on the Pdx1 C-terminal region that has thus far eluded structural characterization. Here we use fluorescence spectrophotometry and protein chemistry to demonstrate that the Pdx1 C terminus is indispensable for PLP synthase activity and mediates intersubunit cross-talk within the enzyme complex. We provide evidence that the C terminus can act as a flexible lid, bridging as well as shielding the active site of an adjacent protomer in Pdx1. We show that ribose 5-phosphate binding triggers strong cooperativity in Pdx1, and the affinity for this substrate is substantially enhanced upon interaction with the Michaelis complex of Pdx2 and glutamine.

Keywords
  • Bacillus subtilis/enzymology
  • Bacterial Proteins/chemistry/metabolism
  • Catalytic Domain/physiology
  • Glutaminase/chemistry/metabolism
  • Glyceraldehyde 3-Phosphate/chemistry/metabolism
  • Ligases/chemistry/metabolism
  • Multienzyme Complexes/chemistry/metabolism
  • Protein Binding/physiology
  • Pyridoxal Phosphate/biosynthesis/chemistry
  • Ribosemonophosphates/chemistry/metabolism
  • Spectrometry
  • Fluorescence
  • Thermotoga maritima/enzymology
  • Transaminases/chemistry/metabolism
  • Xylose/analogs & derivatives/chemistry/metabolism
Citation (ISO format)
RASCHLE, Thomas et al. Intersubunit cross-talk in pyridoxal 5′-phosphate synthase, coordinated by the C terminus of the synthase subunit. In: The Journal of biological chemistry, 2009, vol. 284, n° 12, p. 7706–7718. doi: 10.1074/jbc.M804728200
Main files (1)
Article (Published version)
accessLevelRestricted
Identifiers
ISSN of the journal0021-9258
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