Purification, cloning, and synthesis of a novel salivary anti-thrombin from the mosquito Anopheles albimanus.
An anti-thrombin peptide (anophelin) was isolated from the salivary glands of the mosquito Anopheles albimanus through molecular sieving and reverse-phase high-performance liquid chromatography. The purified peptide inhibited thrombin-induced platelet aggregation, thrombin esterolytic activity on a synthetic substrate, and thrombin cleavage of fibrinogen. The purified anti-thrombin had a molecular mass of 6342.4 Da. Its amino terminus was blocked, but internal sequence yielded three peptide sequences, which were used to design oligonucleotide probes for polymerase chain reaction amplification of salivary gland cDNA and isolation of the full-length clone. Analysis of the sequence of anophelin shows no similarities to any other anti-thrombin peptides. Anophelin was successfully synthesized and characterized to be a tight-binding, specific, and novel inhibitor of thrombin.
Valenzuela
J G
JG
Section of Medical Entomology, Laboratory of Parasitic Diseases, National Institute of Allergy and Infectious Diseases, Bethesda, Maryland 20892-0425, USA.
Francischetti
I M
IM
Ribeiro
J M
JM
D000595:Amino Acid Sequence; D000818:Animals; D000852:Anopheles / Q000737:chemistry*; D001483:Base Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000302:isolation & purification; D005260:Female; D006801:Humans; D019476:Insect Proteins / Q000138:chemical synthesis / Q000235:genetics* / Q000302:isolation & purification* / Q000502:physiology; D008969:Molecular Sequence Data; D010974:Platelet Aggregation / Q000187:drug effects; D010975:Platelet Aggregation Inhibitors / Q000138:chemical synthesis / Q000302:isolation & purification / Q000494:pharmacology; D012469:Salivary Glands / Q000737:chemistry; D012471:Salivary Proteins and Peptides / Q000138:chemical synthesis / Q000235:genetics* / Q000302:isolation & purification* / Q000502:physiology; D013917:Thrombin / Q000037:antagonists & inhibitors* / Q000494:pharmacology
D018076:DNA, Complementary; D019476:Insect Proteins; D010975:Platelet Aggregation Inhibitors; D012471:Salivary Proteins and Peptides; C121275:anophelin protein, Anopheles albimanus; D013917:Thrombin