Tribute to R. G. Boutilier: evidence of a high activity carbonic anhydrase isozyme in the red blood cells of an ancient vertebrate, the sea lamprey Petromyzon marinus.
Carbonic anhydrase (CA) is a multi-functional enzyme that catalyzes the hydration/dehydration of carbon dioxide. In the red blood cell (rbc), CA is necessary to facilitate the transport of carbon dioxide out of the body. Results from earlier biochemical studies indicate that ancient vertebrates, such as agnathans, possess a low activity rbc CA isozyme, whereas more recently evolved vertebrates, such as teleost fish, possess a high activity isozyme. At present, however, the changes in the molecular structure that have resulted in this large increase in catalytic efficiency are unknown. The objective of the current study was therefore to determine the molecular structure of rbc CA in lampreys and compare it to that of teleosts in an effort to ascertain how this important enzyme became more efficient over evolutionary time. Isolation and sequencing of cytoplasmic CA from rbc and gill showed only a single isozyme of 789 bp (262 amino acids). This isozyme was also found in brain and kidney, with no evidence of additional cytoplasmic CA isozymes in other tissues. Phylogenetic analysis grouped this isozyme closely to vertebrate CA VII, which is ancestral to the rbc isozymes in other vertebrates. Interestingly, active site analysis revealed a structure similar to high activity isozymes. A comparative kinetic analysis of CA from rbc lysates and CA fusion proteins showed that the traditional method of determining the turnover number may not be appropriate for all vertebrate CAs. In contrast to previous evidence, lamprey CA was found to be a high activity isozyme. These results suggest that the critical functional characteristics of rbc CA have been highly conserved throughout vertebrate evolution.
Department of Biology, Queen's University, Kingston, Ontario, K7L 3N6, Canada. esbaugha@biology.queensu.ca
A J
AJ
Esbaugh
B L
BL
Tufts
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D002256:Carbonic Anhydrases / Q000097:blood* / Q000235:genetics; D018076:DNA, Complementary / Q000737:chemistry / Q000302:isolation & purification; D004912:Erythrocytes / Q000201:enzymology*; D005880:Gills / Q000201:enzymology; D006801:Humans; D007527:Isoenzymes / Q000097:blood* / Q000235:genetics; D007700:Kinetics; D008969:Molecular Sequence Data; D048428:Petromyzon / Q000097:blood*; D010802:Phylogeny; D016415:Sequence Alignment; D017421:Sequence Analysis; D014018:Tissue Distribution
D018076:DNA, Complementary; D007527:Isoenzymes; D002256:Carbonic Anhydrases