Human Δ32-enoyl-CoA isomerase, type 2: A structural enzymology study on the catalytic role of its ACBP domain and helix-10

Goodluck U. Onwukwe, Petri Kursula, M. Kristian Koski, Werner Schmitz, Rik K. Wierenga

Research output: Contribution to journalArticleResearchpeer-review

11 Citations (Scopus)

Abstract

The catalytic domain of the trimeric human Δ32-enoyl-CoA isomerase, type 2 (HsECI2), has the typical crotonase fold. In the active site of this fold two main chain NH groups form an oxyanion hole for binding the thioester oxygen of the 3E- or 3Z-enoyl-CoA substrate molecules. A catalytic glutamate is essential for the proton transfer between the substrate C2 and C4 atoms for forming the product 2E-enoyl-CoA, which is a key intermediate in the β-oxidation pathway. The active site is covered by the C-terminal helix-10. In HsECI2, the isomerase domain is extended at its N terminus by an acyl-CoA binding protein (ACBP) domain. Small angle X-ray scattering analysis of HsECI2 shows that the ACBP domain protrudes out of the central isomerase trimer. X-ray crystallography of the isomerase domain trimer identifies the active site geometry. A tunnel, shaped by loop-2 and extending from the catalytic site to bulk solvent, suggests a likely mode of binding of the fatty acyl chains. Calorimetry data show that the separately expressed ACBP and isomerase domains bind tightly to fatty acyl-CoA molecules. The truncated isomerase variant (without ACBP domain) has significant enoyl-CoA isomerase activity; however, the full-length isomerase is more efficient. Structural enzymological studies of helix-10 variants show the importance of this helix for efficient catalysis.

Original languageEnglish
Pages (from-to)746-768
Number of pages23
JournalFEBS Journal
Volume282
Issue number4
DOIs
Publication statusPublished - Feb 2015
Externally publishedYes

Keywords

  • crotonase
  • ECI
  • helix-10
  • oxyanion hole
  • SAXS

Cite this