Structure-function relationships of the liver and muscle isoforms of carnitine palmitoyltransferase I.
Elucidation of the membrane topology of carnitine palmitoyltransferase (CPT) I showed that the extreme N-terminus is involved in determining the sensitivity of the liver (L) isoform to malonyl-CoA and suggested that interaction between the two cytosolic segments of the CPT I molecule determines the kinetic characteristics of the enzyme. Work with chimaeric liver/muscle-isoform (L/M) proteins constructed from all six possible combinations of three domains [N-terminus plus transmembrane domain 1 (TM1), loop plus TM2 and C-domain] expressed in Pichia pastoris showed that the precise N-C and TM1-TM2 pairings determine the overall kinetic parameters of the protein. Discrete short sequences within the respective N-terminal regions have negative or positive effects on malonyl-CoA sensitivity (L-isoform) or the K(m) for carnitine (M-isoform) in the full-length proteins, thus imparting to them their distinctive kinetic characteristics. Interactions within N-terminal domains also seem to be important in the targeting of the protein to microsomes in the P. pastoris expression system.
Zammit
V A
VA
Cell Biochemistry, Hannah Research Institute, Ayr KA6 5HL, Scotland, U.K. zammit@main.hri.sari.ac.uk
Price
N T
NT
Fraser
F
F
Jackson
V N
VN
D000818:Animals; D002334:Carnitine O-Palmitoyltransferase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D020134:Catalytic Domain; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D008316:Malonyl Coenzyme A / Q000378:metabolism; D009132:Muscles / Q000201:enzymology*; D010843:Pichia / Q000235:genetics; D017434:Protein Structure, Tertiary; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D017384:Sequence Deletion / Q000235:genetics; D013329:Structure-Activity Relationship
D007527:Isoenzymes; D011993:Recombinant Fusion Proteins; D008316:Malonyl Coenzyme A; D002334:Carnitine O-Palmitoyltransferase