Loop closure and intersubunit communication in tryptophan synthase

Biochemistry. 1998 Apr 21;37(16):5394-406. doi: 10.1021/bi9728957.

Abstract

Crystal structures of wild-type tryptophan synthase alpha2beta2 complexes from Salmonella typhimurium were determined to investigate the mechanism of allosteric activation of the alpha-reaction by the aminoacrylate intermediate formed at the beta-active site. Using a flow cell, the aminoacrylate (A-A) intermediate of the beta-reaction () was generated in the crystal under steady state conditions in the presence of serine and the alpha-site inhibitor 5-fluoroindole propanol phosphate (F-IPP). A model for the conformation of the Schiff base between the aminoacrylate and the beta-subunit cofactor pyridoxal phosphate (PLP) is presented. The structure is compared with structures of the enzyme determined in the absence (TRPS) and presence (TRPSF-IPP) of F-IPP. A detailed model for binding of F-IPP to the alpha-subunit is presented. In contrast to findings by Hyde et al. [(1988) J. Biol. Chem. 263,17857-17871] and Rhee et al. [(1997) Biochemistry 36, 7664-7680], we find that the presence of an alpha-site alone ligand is sufficient for loop alphaL6 closure atop the alpha-active site. Part of this loop, alphaThr183, is important not only for positioning the catalytic alphaAsp60 but also for coordinating the concomitant ordering of loop alphaL2 upon F-IPP binding. On the basis of the three structures, a pathway for communication between the alpha- and beta-active sites has been established. The central element of this pathway is a newly defined rigid, but movable, domain that on one side interacts with the alpha-subunit via loop alphaL2 and on the other side with the beta-active site. These findings provide a structural basis for understanding the allosteric properties of tryptophan synthase.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Alanine / analogs & derivatives
  • Alanine / metabolism
  • Allosteric Site
  • Crystallography, X-Ray
  • Enzyme Activation
  • Ligands
  • Models, Molecular
  • Protein Structure, Secondary
  • Protein Structure, Tertiary
  • Salmonella typhimurium / enzymology
  • Serine / metabolism
  • Structure-Activity Relationship
  • Tryptophan Synthase / chemistry*

Substances

  • Ligands
  • Serine
  • dehydroalanine
  • Tryptophan Synthase
  • Alanine

Associated data

  • PDB/1A50
  • PDB/1A5S
  • PDB/2WSY