A novel family of anticoagulants from the saliva of Ixodes scapularis.
Using biochemical and molecular approaches, we have identified a 9.8 kDa protein in the saliva of Ixodes scapularis that inhibits the intrinsic pathway of coagulation. The 9.8 kDa anticoagulant protein was purified by reverse-phase HPLC and its N-terminal amino acid sequence determined. The N-terminal sequence showed homology with Salp14, an immuno-dominant antigen present in the saliva of engorging I. scapularis nymphs. Recombinant Salp14 expressed in Escherichia coli prolonged the activated partial thromboplastin time (APTT) of human plasma in a dose-dependent manner and was a specific inhibitor of factor Xa. A cDNA encoding a 9.3 kDa protein, Salp9Pac, was subsequently isolated from an I. scapularis salivary gland cDNA library. Salp9Pac showed 93% identity to the N-terminal sequence of the anticoagulant purified by HPLC. These data indicate that the anticoagulant protein purified by HPLC, Salp9Pac and Salp14 are members of a family of novel coagulation protease inhibitors present in tick saliva. While recombinant Salp9Pac did not show biological activity in the assays tested currently, it is likely to be mechanistically different from its paralogues. This raises the possibility that ticks may enhance their adaptive ability to cope with a wide spectrum of proteases, by transcribing such structurally related anticoagulant proteins with different functions.
Narasimhan
S
S
Department of Internal Medicine, Yale University School of Medicine, New Haven, Connecticut 06520-8031, USA.
Koski
R A
RA
Beaulieu
B
B
Anderson
J F
JF
Ramamoorthi
N
N
Kantor
F
F
Cappello
M
M
Fikrig
E
E
D000595:Amino Acid Sequence; D000818:Animals; D000925:Anticoagulants / Q000737:chemistry / Q000302:isolation & purification*; D003001:Cloning, Molecular; D017931:DNA Primers; D004926:Escherichia coli / Q000235:genetics; D015723:Gene Library; D019476:Insect Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018884:Ixodes / Q000502:physiology*; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry; D012463:Saliva / Q000378:metabolism*; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid
D000925:Anticoagulants; D017931:DNA Primers; D019476:Insect Proteins; D011994:Recombinant Proteins