Purification and structural determination of a phosphorylated peptide with anti-calcification and chitin-binding activities in the exoskeleton of the crayfish, Procambarus clarkii.
Organic matrices in calcified hard tissues have been considered to control calcification. A matrix peptide, designated CAP-1, was extracted and purified by anion-exchange and reverse-phase high performance liquid chromatographies from the exoskeleton of the crayfish, Procambarus clarkii. The amino acid sequence of CAP-1 was determined by mass spectral and sequence analyses of the intact peptide and its enzymatically digested peptides. CAP-1 consisted of 78 amino acid residues, including a phosphoserine residue, and was rich in acidic amino acid residues. CAP-1 had a Rebers-Riddiford consensus sequence, which is conserved in cuticle proteins from many arthropods. CAP-1 inhibited precipitation of calcium carbonate in an in vitro anticalcification assay dose-dependently, and completely inhibited it at 3 x 10(-7) M. CAP-1 also showed chitin-binding ability, indicating that this molecule was bifunctional and played an important role in formation of the exoskeleton.
Inoue
H
H
Department of Applied Biological Chemistry, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Bunkyo, Japan.
Ozaki
N
N
Nagasawa
H
H
D000595:Amino Acid Sequence; D000818:Animals; D003400:Astacoidea / Q000737:chemistry*; D002114:Calcinosis / Q000517:prevention & control*; D002119:Calcium Carbonate / Q000737:chemistry; D002686:Chitin / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D006868:Hydrolysis; D007202:Indicators and Reagents; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry* / Q000302:isolation & purification / Q000494:pharmacology* / Q000502:physiology*; D010766:Phosphorylation; D010768:Phosphoserine / Q000737:chemistry; D011485:Protein Binding; D013056:Spectrophotometry, Ultraviolet
D007202:Indicators and Reagents; D010455:Peptides; C448763:calcification-associated peptide-1, crayfish; D002686:Chitin; D010768:Phosphoserine; D002119:Calcium Carbonate