Close similarity of baculovirus-expressed n-chimaerin and protein kinase C alpha as phorbol ester receptors.
n-Chimaerin is a recently described phorbol ester receptor that shares homology in its N-terminal region with the cysteine-rich zinc finger domain of protein kinase C. We have expressed n-chimaerin in insect cells using the baculovirus system and have used the isolated, recombinant n-chimaerin to characterize phorbol ester binding and structure-activity relations, lipid requirements, and inhibitor sensitivity. We find that n-chimaerin expressed in the baculovirus system bound [3H]phorbol 12,13-dibutyrate with high affinity (0.17 +/- 0.01 nM). Although having only a single cysteine-rich zinc finger region compared to two for protein kinase C, n-chimaerin thus closely resembled protein kinase C alpha. n-Chimaerin was likewise virtually indistinguishable from protein kinase C alpha in phorbol ester structure-activity relations, in phospholipid requirements, and in inhibition of binding by sphingosine and calphostin C, protein kinase C inhibitors acting on the regulatory domain. We conclude that a number of typical approaches used to implicate protein kinase C in biological function in cells do not discriminate between the n-chimaerin and protein kinase C classes of phorbol ester receptors.
Areces
L B
LB
Molecular Mechanisms of Tumor Promotion Section, NCI, National Institutes of Health, Bethesda, Maryland 20892.
Kazanietz
M G
MG
Blumberg
P M
PM
D000595:Amino Acid Sequence; D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D001483:Base Sequence; D029742:Caenorhabditis elegans Proteins; D002352:Carrier Proteins; D002478:Cells, Cultured; D020848:Chimerin 1; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D009036:Moths / Q000166:cytology; D009281:Naphthalenes; D009419:Nerve Tissue Proteins / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D015240:Phorbol 12,13-Dibutyrate / Q000378:metabolism; D010718:Phosphatidylserines / Q000494:pharmacology; D010743:Phospholipids / Q000494:pharmacology; D011083:Polycyclic Compounds / Q000494:pharmacology; D011493:Protein Kinase C / Q000235:genetics / Q000378:metabolism*; D051571:Protein Kinase C-alpha; D011955:Receptors, Drug / Q000378:metabolism*; D013110:Sphingosine / Q000494:pharmacology
D029742:Caenorhabditis elegans Proteins; D002352:Carrier Proteins; D020848:Chimerin 1; D007527:Isoenzymes; D009281:Naphthalenes; D009419:Nerve Tissue Proteins; D010718:Phosphatidylserines; D010743:Phospholipids; D011083:Polycyclic Compounds; D011955:Receptors, Drug; C083141:phorbol ester binding protein; C027776:phorbol ester receptor; D015240:Phorbol 12,13-Dibutyrate; D011493:Protein Kinase C; D051571:Protein Kinase C-alpha; C058819:calphostin C; D013110:Sphingosine