Anticoagulant of blood sucking insect salivary glands

我们分析了血吸虫Rhodnius prolixus唾液腺中的生理活性物质。通过对活化部分凝血活酶时间(APTT)和凝血酶原时间(PT)的分析，发现红景天唾液腺提取物具有两种不同的抗凝血活性。我们的分析表明凝血酶在凝血级联中既有内在途径、Xase复合体和共同途径的抑制活性，也有凝血酶(因子11a)的抑制活性。我们重点分析了Xase抑制剂，并试图进行纯化和鉴定。该抑制物是一种分子量约为20 kDa的血红蛋白。我们将其命名为普罗利辛-S。普罗利辛-S具有抑制凝血因子X激活的活性，即Xase复合体，包括凝血因子IX、VIII、磷脂和钙。我们正在研究Xase复合体对普罗利辛-S的封闭机制。用普罗利辛-S多克隆抗体从红景天唾液腺文库中克隆了普罗利辛-S的全长基因，并进行了序列测定。利用杆状病毒昆虫细胞表达系统表达了普罗利欣-S，并进行了层析纯化。重组普罗利欣-S的抗凝血活性几乎与天然普罗利辛相当。普罗利辛-S很容易与亚铁血红素重组为血红素蛋白，并能可逆地与一氧化氮(NO)结合。游离普罗利欣-S对兔血管平滑肌有扩张作用。这种分子名为普罗利欣-S，是一种独特的蛋白质，既有抗凝血活性，又有血管扩张活性。

We have analyzed physiologically active compounds from the salivary glands of blood suckinh insect, Rhodnius prolixus. We found two different anti-coagulant activities from the extract of Rhodnius salivary glands by analyzing activated partial thromboplastin time (APTT) and prothrombin time (PT). Our analyzes showed inhibitory activities of both intrinsic pathway, Xase complex and common pathway, thrombin (factor lla) in the coagulant cascade.We focussed our analyzes on the Xase inhibitor, and tried to purify and characterize. This inhibitor is a hemeprotein with molecular weight of about 20 kDa. We named it Prolixin-S.Prolixin-S has an inhibitory activity of factor X activation, i.e.Xase complex, which include factor IX,VIII,phospholipid and Ca. We are studying on the mechanism of Xase complex blocking with Prolixin-S.Prolixin-S cDNA was cloned from Rhodnius salivary gland cDNA library with polyclonal antibody to Prolixin-S,and sequenced. Prolixin-S was expressed by baculovirus insect cell expression system and purified chromatographically. Recominant Prolixin-S had anticoagulant activity as almost same level as native Prolixin-S.rProlixin-S was easily reconstituted as heme-protein with hemine and can be bound nitric oxide (NO) reversiblly. NO bound prolixin-S has a vasodilation activity to rabbit vascular smooth muscle. This molecule, Prolixin-S,is a unique protein which has both anti-coagulant activity and vasodilating activity.