Packing of transmembrane domain 2 of carnitine palmitoyltransferase-1A affects oligomerization and malonyl-CoA sensitivity of the mitochondrial outer membrane protein.
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 IC(50) 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.[1]References
- Packing of transmembrane domain 2 of carnitine palmitoyltransferase-1A affects oligomerization and malonyl-CoA sensitivity of the mitochondrial outer membrane protein. Jenei, Z.A., Warren, G.Z., Hasan, M., Zammit, V.A., Dixon, A.M. FASEB J. (2011) [Pubmed]
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