Kiwi protein inhibitor of pectin methylesterase amino-acid sequence and structural importance of two disulfide bridges.
A protein acting as a powerful inhibitor of plant pectin methylesterase was isolated from kiwi (Actinidia chinensis) fruit. The complete amino-acid sequence of the pectin methylesterase inhibitor (PMEI) was determined by direct protein analysis. The sequence comprises 152 amino-acid residues, accounting for a molecular mass of 16 277 Da. The far-UV CD spectrum indicated a predominant alpha-helix conformation in the secondary structure. The protein has five cysteine residues but neither tryptophan nor methionine. Analysis of fragments obtained after digestion of the protein alkylated without previous reduction identified two disulfide bridges connecting Cys9 with Cys18, and Cys74 with Cys114; Cys140 bears a free thiol group. A database search pointed out a similarity between PMEI and plant invertase inhibitors. In particular, the four Cys residues, which in PMEI are involved in the disulfide bridges, are conserved. This allows us to infer that also in the homologous proteins, whose primary structure was deduced only by cDNA sequencing, those cysteine residues are engaged in two disulfide bridges, and constitute a common structural motif. The comparison of the sequence of these inhibitors confirms the existence of a novel class of proteins with moderate but significant sequence conservation, comprising plant proteins acting as inhibitors of sugar metabolism enzymes, and probably involved in various steps of plant development.
Camardella
L
L
Institute of Protein Biochemistry and Enzymology, C.N.R.
Carratore
V
V
Ciardiello
M A
MA
Servillo
L
L
Balestrieri
C
C
Giovane
A
A
D000595:Amino Acid Sequence; D001224:Aspartic Acid / Q000494:pharmacology; D002265:Carboxylic Ester Hydrolases / Q000037:antagonists & inhibitors*; D002851:Chromatography, High Pressure Liquid; D002942:Circular Dichroism; D003545:Cysteine / Q000737:chemistry; D018076:DNA, Complementary / Q000378:metabolism; D004220:Disulfides; D005638:Fruit / Q000737:chemistry*; D008969:Molecular Sequence Data; D010940:Plant Proteins / Q000737:chemistry* / Q000302:isolation & purification; D017433:Protein Structure, Secondary; D020539:Sequence Analysis, Protein; D017386:Sequence Homology, Amino Acid
D018076:DNA, Complementary; D004220:Disulfides; C412789:PMEI protein, Actinidia chinensis; D010940:Plant Proteins; D001224:Aspartic Acid; D002265:Carboxylic Ester Hydrolases; C022323:pectinesterase; D003545:Cysteine