Jenei, Z. A., Warren, Gemma, Hasan, M., Zammit, V. A. (Victor A.) and Dixon, Ann M.. (2011) Packing of transmembrane domain 2 of carnitine palmitoyltransferase-1A affects oligomerization and malonyl-CoA sensitivity of the mitochondrial outer membrane protein. FASEB Journal, Vol.25 (No.12). pp. 4522-4530. ISSN 0892-6638

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Abstract

The purpose of this study was to investigate the sequence-dependence of oligomerization of transmembrane domain 2 (TM2) of rat carnitine palmitoyltransferase 1A (rCPT1A), to elucidate the role of this domain in the function of the full-length enzyme. Oligomerization of TM2 was studied qualitatively using complementary genetic assays that facilitate measurement of helix-helix interactions in the Escherichia coli inner membrane, and multiple quantitative biophysical methods. The effects of TM2-mutations on oligomerization and malonyl-CoA inhibition of the full-length enzyme (expressed in the yeast Pichia pastoris) were quantified. Changes designed to disrupt close-packing of the GXXXG(A) motifs reduced the oligomeric state of the corresponding TM2 peptides from hexamer to trimer (or lower), a reduction also observed on mutation of the TM2 sequence in the full-length enzyme. Disruption of these GXXXG(A) motifs had a parallel effect on the malonyl-CoA sensitivity of rCPT1A, reducing the IC50 from 30.3 ± 5.0 to 3.0 ± 0.6 μM. For all measurements, wild-type rCPT1A was used as a control alongside various appropriate (e.g., molecular mass) standards. Our results suggest that sequence-determined, TM2-mediated oligomerization is likely to be involved in the modulation of malonyl-CoA inhibition of CPT1A in response to short- and long-term changes in protein-protein and protein-lipid interactions that occur in vivo. —Jenei, Z. A., Warren, G. Z. L., Hasan, M., Zammit, V. A., Dixon, A. M. Packing of transmembrane domain 2 of carnitine palmitoyltransferase-1A affects oligomerization and malonyl-CoA sensitivity of the mitochondrial outer membrane protein.

Item Type:	Journal Article
Subjects:	Q Science > QD Chemistry Q Science > QP Physiology R Medicine > RC Internal medicine
Divisions:	Faculty of Science > Chemistry Faculty of Medicine > Warwick Medical School > Translational & Systems Medicine > Metabolic and Vascular Health Faculty of Medicine > Warwick Medical School
Library of Congress Subject Headings (LCSH):	Fatty acids -- Metabolism, Diabetes -- Pathophysiology, Ultracentrifugation, Membrane proteins
Journal or Publication Title:	FASEB Journal
Publisher:	Federation of American Societies for Experimental Biology
ISSN:	0892-6638
Official Date:	December 2011
Volume:	Vol.25
Number:	No.12
Page Range:	pp. 4522-4530
Identification Number:	10.1096/fj.11-192005
Status:	Peer Reviewed
Publication Status:	Published
Access rights to Published version:	Restricted or Subscription Access
URI:	http://wrap.warwick.ac.uk/id/eprint/40211

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