Sugimoto M, Nakajima N
Research Institute for Bioresources, Okayama University, Kurashiki, Japan.
Biosci Biotechnol Biochem. 2001 Jul;65(7):1575-80. doi: 10.1271/bbb.65.1575.
An earthworm, Lumbricus rubellus, produces alkaline trypsin-like proteases that are greater than trypsins in their stability and strong tolerance to organic solvents. cDNAs encoding strong fibrinolytic proteases (F-III-2 and F-III-1) in the six isozymes were cloned and sequenced to study their stability-structure relationship. The cDNAs of F-III-2 and F-III-1 comprised 1011 and 973 bp and included open reading frames that encode polypeptides of 245 and 246 amino acid residues, respectively. The deduced amino acid sequences of F-III-2 and F-III-1 have 7 and 8 activation peptides in the N-termini respectively, indicating that they are translated as proenzymes and modified to active forms by posttranslational processing. They showed similarity to mammalian serine proteases and conserved the catalytic amino acid residues, however, neither arginine nor lysine residues were present in the autolysis region. The gene encoding the native form of F-III-2 was expressed in Pichia pastoris to produce and secrete the earthworm protease in the culture medium, which dissolves an artificial fibrin plate.
赤子爱胜蚓会产生碱性胰蛋白酶样蛋白酶，其稳定性高于胰蛋白酶，且对有机溶剂具有很强的耐受性。对六种同工酶中编码强纤维蛋白溶解蛋白酶（F-III-2和F-III-1）的cDNA进行了克隆和测序，以研究它们的稳定性与结构的关系。F-III-2和F-III-1的cDNA分别由1011和973个碱基对组成，包含分别编码245和246个氨基酸残基的开放阅读框。F-III-2和F-III-1推导的氨基酸序列在N端分别有7个和8个激活肽，这表明它们作为酶原被翻译，并通过翻译后加工修饰为活性形式。它们与哺乳动物丝氨酸蛋白酶具有相似性，并保留了催化氨基酸残基，然而，自溶区域中既没有精氨酸也没有赖氨酸残基。编码F-III-2天然形式的基因在毕赤酵母中表达，以便在培养基中产生并分泌蚯蚓蛋白酶，该酶可溶解人工纤维蛋白平板。
