Meeting Abstract

40.1  Wednesday, Jan. 5  Creatine Kinase is Present in an Alveolate Protozoan HOFFMAN, GG; SUZUKI, T; ELLINGTON, WR*; Florida St. Univ., Tallahassee; Kochi Univ., Kochi, Japan; Florida St. Univ., Tallahassee wellington@admin.fsu.edu

Creatine kinase (CK) is present in animals as a family of isoforms – cytoplasmic, mitochondrial and flagellar- each targeted to different intracellular compartments. Recently deposited genomic and EST sequences show that the aveolate protozoan Perkinsus marinus has at least two CK-like genes. One codes for a protein with similarity to cytoplasmic and mitochondrial CKs. The other gene codes for a protein consisting of two fused CK domains similar to the contiguous trimeric flagellar CKs. We used RTPCR to obtain the cDNA for one domain of the contiguous dimeric CK. The cDNA was ligated into an expression vector which was used to transform DE3 E. coli cells. Subsequent expression, purification and assay of the recombinant protein showed that this cDNA codes for a true CK. The specific activity of the purified P. marinus CK was in the same range as has been observed for typical CKs from animals. P. marinus has a complex life cycle involving a motile zoospore stage. Zoospores have two long flagella thereby creating reaction-diffusion constraints for energy transport between the ATP source (mitochondria) and distal ATP sinks (dynein ATPases). The CK system in P. marinus is likely involved in mitigating spatial mismatches of ATP supply and demand. The presence of CK in the two major eukaryotic lineages (bikont, unikont) would suggest that this enzyme evolved at the dawn of the radiation of eukaryotes. This event may have been associated with the transition from a benthic crawling or gliding lifestyle to the planktonic condition wherein motility was powered by ciliary/flagellar elements. (Supported by NSF grant IOS-0542236 to WRE).