Cloning and characterization of trypsin- and chymotrypsin-like proteases from the midgut of the sand fly vector Phlebotomus papatasi.
Trypsin and chymotrypsin serine proteases are the main digestive proteases in Diptera midguts and are also involved in many aspects of the vector-parasite relationship. In sand flies, these proteases have been shown to be a potential barrier to Leishmania growth and development within the midgut. Here we describe the sequence and partial characterization of six Phlebotomus papatasi midgut serine proteases: two chymotrypsin-like (Ppchym1 and Ppchym2) and four trypsin-like (Pptryp1-Pptryp4). All six enzymes show structural features typical to each type, including the histidine, aspartic acid, and serine (H/D/S) catalytic triad, six conserved cysteine residues, and other amino acid residues involved in substrate specificity. They also show a high degree of homology (40-60% identical residues) with their counterparts from other insect vectors, such as Anopheles gambiae and Aedes aegypti. The mRNA expression profiles of these six proteases vary considerably: two trypsin-like proteases (Pptryp1 and Pptryp2) are downregulated and one (Pptryp4) upregulated upon blood feeding. The two chymotrypsin-like enzymes display expression behavior similar to that of the early and late trypsins from Ae. aegypti.
Ramalho-Ortigão
J M
JM
Intracellular Parasite Biology Section, Laboratory of Parasitic Diseases, National Institute of Allergy and Infectious Diseases, 4 Center Drive, Building 4, Room 126, MSC-0425, NIH, Bethesda, MD 20892-0425, USA.
Kamhawi
S
S
Rowton
E D
ED
Ribeiro
J M C
JM
Valenzuela
J G
JG
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002918:Chymotrypsin / Q000235:genetics*; D017931:DNA Primers; D004064:Digestive System / Q000201:enzymology*; D015971:Gene Expression Regulation, Enzymologic; D008969:Molecular Sequence Data; D010691:Phlebotomus / Q000145:classification / Q000201:enzymology*; D010802:Phylogeny; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D012697:Serine Endopeptidases / Q000235:genetics*; D014357:Trypsin / Q000235:genetics*
D000596:Amino Acids; D017931:DNA Primers; D012697:Serine Endopeptidases; D002918:Chymotrypsin; D014357:Trypsin