VARIANT CDNA SEQUENCES OF HUMAN ATP-CITRATE LYASE - CLONING, EXPRESSION, AND PURIFICATION FROM BACULOVIRUS-INFECTED INSECT CELLS

ATP:citrate lyase (ACL) is a major generator of cytosolic acetyl-coenz yme A, which is required for both fatty acid and cholesterol biosynthe sis, The human ACL (hACL) cDNA was cloned by RT-PCR, and our results i ndicate the existence of previously unknown sequence variations in hAC L. Expression of the hACL cDNA in Spodoptera frugiperda 9 insect cells resulted in the production of high levels of soluble, active enzyme, The recombinant protein (re-hACL) was purified to homogeneity from the soluble lysate of infected cells and was observed to exist as a tetra mer by gel filtration chromatography. Kinetic analyses indicated that the re-hACL and rat ACL have very similar enzymological properties. Th e facile preparation of milligram quantities of purified, active re-hA CL affords the opportunity to characterize the enzyme for structure-ba sed design of hypolipidemic drugs, and to further examine the function al significance of the sequence variations. (C) 1997 Academic Press.