Conantokin-G precursor and its role in gamma-carboxylation by a vitamin K-dependent carboxylase from a Conus snail.
Conantokin-G isolated from the marine snail Conus geographus is a 17-amino acid gamma-carboxyglutamate (Gla)-containing peptide that inhibits the N-methyl-D-aspartate receptor. We describe the cloning and sequence of conantokin-G cDNA and the possible role of the propeptide sequence. The cDNA encodes a 100amino acid peptide. The N-terminal 80 amino acids constitute the prepro-sequence, and the mature peptide is derived from the remaining C-terminal residues after proteolysis, C-terminal amidation, and a unique post-translational modification, gamma-carboxylation of glutamate residues to Gla. Mature conantokin-G peptide containing Glu residues (E.Con-G) in place of Gla is a poor substrate for the vitamin K-dependent gamma-glutamyl carboxylase (apparent Km = 3.4 mM). Using peptides corresponding to different segments of the propeptide we investigated a potential role for the propeptide sequences in gamma-carboxylation. Propeptide segment -20 to -1 covalently linked to E.Con-G or the synthetic pentapeptide FLEEL increased their apparent affinities 2 orders of magnitude. These substrates are not efficiently carboxylated by the bovine microsomal gamma-glutamyl carboxylase, suggesting differences in specificities between the Conus and the mammalian enzyme. However, the role of propeptide in enhancing the efficiency of carboxylation is maintained.
Bandyopadhyay
P K
PK
Department of Biology, University of Utah, Salt Lake City, Utah 84112, USA. bandyop@biology.utah.edu
Colledge
C J
CJ
Walker
C S
CS
Zhou
L M
LM
Hillyard
D R
DR
Olivera
B M
BM
D015055:1-Carboxyglutamic Acid / Q000096:biosynthesis; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D019735:Carbon-Carbon Ligases / Q000378:metabolism*; D002417:Cattle; D003001:Cloning, Molecular; D020916:Conotoxins; D018691:Excitatory Amino Acid Antagonists / Q000737:chemistry / Q000494:pharmacology; D018698:Glutamic Acid / Q000378:metabolism; D007700:Kinetics; D008861:Microsomes / Q000201:enzymology; D008969:Molecular Sequence Data; D008978:Mollusk Venoms / Q000737:chemistry* / Q000633:toxicity; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D010456:Peptides, Cyclic / Q000737:chemistry* / Q000633:toxicity; D011498:Protein Precursors / Q000737:chemistry* / Q000502:physiology; D011499:Protein Processing, Post-Translational / Q000502:physiology; D017422:Sequence Analysis, DNA; D012908:Snails / Q000201:enzymology; D013379:Substrate Specificity
D020916:Conotoxins; D018691:Excitatory Amino Acid Antagonists; D008978:Mollusk Venoms; D010446:Peptide Fragments; D010456:Peptides, Cyclic; D011498:Protein Precursors; D018698:Glutamic Acid; D015055:1-Carboxyglutamic Acid; C055407:conotoxin GV; D019735:Carbon-Carbon Ligases; C020660:glutamyl carboxylase