Dipetalodipin, a novel multifunctional salivary lipocalin that inhibits platelet aggregation, vasoconstriction, and angiogenesis through unique binding specificity for TXA2, PGF2alpha, and 15(S)-HETE.
Dipetalodipin (DPTL) is an 18 kDa protein cloned from salivary glands of the triatomine Dipetalogaster maxima. DPTL belongs to the lipocalin superfamily and has strong sequence similarity to pallidipin, a salivary inhibitor of collagen-induced platelet aggregation. DPTL expressed in Escherichia coli was found to inhibit platelet aggregation by collagen, U-46619, or arachidonic acid without affecting aggregation induced by ADP, convulxin, PMA, and ristocetin. An assay based on incubation of DPTL with small molecules (e.g. prostanoids, leukotrienes, lipids, biogenic amines) followed by chromatography, mass spectrometry, and isothermal titration calorimetry showed that DPTL binds with high affinity to carbocyclic TXA(2), TXA(2) mimetic (U-46619), TXB(2), PGH(2) mimetic (U-51605), PGD(2,) PGJ(2), and PGF(2α). It also interacts with 15(S)-HETE, being the first lipocalin described to date to bind to a derivative of 15-lipoxygenase. Binding was not observed to other prostaglandins (e.g. PGE(1), PGE(2), 8-iso-PGF(2α), prostacyclin), leukotrienes (e.g. LTB(4), LTC(4), LTD(4), LTE(4)), HETEs (e.g. 5(S)-HETE, 12(S)-HETE, 20-HETE), lipids (e.g. arachidonic acid, PAF), and biogenic amines (e.g. ADP, serotonin, epinephrine, norepinephrine, histamine). Consistent with its binding specificity, DPTL prevents contraction of rat uterus stimulated by PGF(2α) and induces relaxation of aorta previously contracted with U-46619. Moreover, it inhibits angiogenesis mediated by 15(S)-HETE and did not enhance inhibition of collagen-induced platelet aggregation by SQ29548 (TXA(2) antagonist) and indomethacin. A 3-D model for DPTL and pallidipin is presented that indicates the presence of a conserved Arg(39) and Gln(135) in the binding pocket of both lipocalins. Results suggest that DPTL blocks platelet aggregation, vasoconstriction, and angiogenesis through binding to distinct eicosanoids involved in inflammation.
Assumpção
Teresa C F
TC
Vector Biology Section, Laboratory of Malaria and Vector Research, NIAID, National Institutes of Health, Bethesda, Maryland 20892-8132, USA.
Alvarenga
Patricia H
PH
Ribeiro
José M C
JM
Andersen
John F
JF
Francischetti
Ivo M B
IM
D000818:Animals; D001011:Aorta / Q000187:drug effects / Q000378:metabolism; D015237:Dinoprost / Q000378:metabolism*; D005260:Female; D006736:Horses; D006893:Hydroxyeicosatetraenoic Acids / Q000737:chemistry*; D019476:Insect Proteins / Q000737:chemistry*; D054834:Lipocalins / Q000737:chemistry / Q000378:metabolism*; D009389:Neovascularization, Pathologic; D010974:Platelet Aggregation / Q000187:drug effects*; D051381:Rats; D017208:Rats, Wistar; D012463:Saliva / Q000378:metabolism*; D012469:Salivary Glands / Q000378:metabolism; D013928:Thromboxane A2 / Q000378:metabolism*; D014225:Triatominae / Q000378:metabolism*; D014599:Uterus / Q000187:drug effects; D014661:Vasoconstriction
D006893:Hydroxyeicosatetraenoic Acids; D019476:Insect Proteins; D054834:Lipocalins; C554827:dipetalodipin protein, Dipetalogaster maxima; D013928:Thromboxane A2; C025984:15-hydroxy-5,8,11,13-eicosatetraenoic acid; D015237:Dinoprost