Isolation, characterization and nucleotide sequence of the muscle isoforms of creatine kinase from the Antarctic teleost Chaenocephalus aceratus.
Creatine kinase (CK) was isolated from the white muscle of the Antarctic icefish Chaenocephalus aceratus, which is deficient in glycolytic capacity. C. aceratus white myotomal creatine kinase (MMCK) displayed an apparent K(m) at 0.5 degrees C of 0.06 mM for ADP and 17 mM for Phosphocreatine. These K(m) values are similar to those reported for other vertebrate MMCKs at their physiologically relevant body temperatures. C. aceratus MMCK exhibited optimal activity at pH of 7.6-7.7 at 0.5 degrees C, in contrast to rabbit MMCK which had optimum activity at pH 6.2 at 30 degrees C. The apparent V(max) of C. aceratus MMCK at 0.5 degrees C is 94+/-4 S.D. (n=9) micromol ATP/min/mg (i.e. U/mg), which is comparable to rabbit MMCK assayed at 20 degrees C and 8-fold greater than rabbit MMCK measured at 0.5 degrees C. DEAE chromatography of C. aceratus white muscle CK resolved two distinct activity peaks. Cloning and sequencing of C. aceratus CK cDNAs confirmed that two muscle-specific isoforms of CK were expressed that were distinct from the mitochondrial and brain isoforms. Icefish MMCK was sensitive to transient temperature elevation, and the DEAE-fractionated forms were highly unstable. These results indicate that C. aceratus MMCK displays significant activity at physiological temperature and intracellular pH of icefish muscle that could contribute to sustaining energy charge during burst-swimming.
Winnard
Paul
P
Department of Biochemistry, Microbiology, and Molecular Biology, School of Marine Sciences, University of Maine, 5735 Hitchner Hall, Orono, ME 04469-5735, USA.
Cashon
Robert E
RE
Sidell
Bruce D
BD
Vayda
Michael E
ME
D000222:Adaptation, Physiological; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D000864:Antarctic Regions; D001483:Base Sequence; D003402:Creatine Kinase / Q000737:chemistry* / Q000302:isolation & purification* / Q000378:metabolism; D005399:Fishes; D006863:Hydrogen-Ion Concentration; D007527:Isoenzymes / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D007700:Kinetics; D008969:Molecular Sequence Data; D018482:Muscle, Skeletal / Q000201:enzymology*; D010725:Phosphocreatine / Q000378:metabolism; D011485:Protein Binding; D011817:Rabbits; D017422:Sequence Analysis, DNA; D013696:Temperature
D007527:Isoenzymes; D010725:Phosphocreatine; D000255:Adenosine Triphosphate; D003402:Creatine Kinase