Characterization of two crustin antimicrobial peptides from the freshwater crayfish Pacifastacus leniusculus.
The two bacteria-induced crustin genes, Plcrustin1 and Plcrustin2, previously found in the hemocyte cDNA library of Pacifastacus leniusculus, contain the open reading frames of 357 bp encoding a putative protein of 118 amino acid residues and 330 bp encoding a putative protein of 109 amino acid residues, respectively. The carboxyl-terminal part of the two crustins possesses, respectively, 7 and 8 conserved cysteine residues representation of a WAP domain that is found in carcinins and crustins in other several crustaceans. The amino acid sequences of Plcrustin1 and Plcrustin2 show that they belong to type I crustins. In order to characterize their properties and biological activities, the two recombinant crustin proteins were produced in the Escherichia coli expression system. Antimicrobial assays showed that the growth of only one Gram-positive bacterium, Micrococcus luteus M1 11, was inhibited by the recombinant Plcrustin1 and Plcrustin2 with MIC of about 0.07-0.27 microM and 3.5-8 microM, respectively. In addition, the study of inhibition mechanism revealed that the antimicrobial activity of the two recombinant crustin proteins was a result of bactericidal effect. However, the two crustins did not exhibit the inhibitory activities against trypsin, chymotrypsin, elastase and subtilisin A.
Donpudsa
Suchao
S
Department of Comparative Physiology, Uppsala University, Norbyvägen 18 A, SE-752 36 Uppsala, Sweden.
Rimphanitchayakit
Vichien
V
Tassanakajon
Anchalee
A
Söderhäll
Irene
I
Söderhäll
Kenneth
K
D000595:Amino Acid Sequence; D000818:Animals; D000890:Anti-Infective Agents / Q000378:metabolism* / Q000633:toxicity; D023181:Antimicrobial Cationic Peptides / Q000378:metabolism* / Q000633:toxicity; D003400:Astacoidea / Q000502:physiology*; D004591:Electrophoresis, Polyacrylamide Gel; D006434:Hemocytes / Q000378:metabolism; D008826:Microbial Sensitivity Tests; D011994:Recombinant Proteins / Q000378:metabolism; D015842:Serine Proteinase Inhibitors / Q000378:metabolism* / Q000633:toxicity
D000890:Anti-Infective Agents; D023181:Antimicrobial Cationic Peptides; D011994:Recombinant Proteins; D015842:Serine Proteinase Inhibitors