Yamagata H, Masuzawa T, Nagaoka Y, Ohnishi T, Iwasaki T
Laboratory of Biochemistry, Faculty of Agriculture, Kobe University, Japan.
J Biol Chem. 1994 Dec 30;269(52):32725-31.
Cucumisin is a thermostable alkaline serine protease that is found in the juice of melon fruits (Cucumis melo L.). We have determined the complete nucleotide sequence of a cucumisin cDNA (2,552 nucleotides) and deduced the corresponding amino acid sequence. The open reading frame of the cDNA consists of 731 codons and encodes a large precursor (molecular weight, 78,815) relative to the observed size of mature native cucumisin (67 kDa). Sequence comparisons reveal that cucumisin has several features in common with the microbial proteases of the subtilisin family. The highly conserved sequences to the proximal regions of the catalytic triad amino acids Asp, His, and Ser, together with the substrate binding site in subtilisin, can be found within the deduced amino acid sequence of the protease domain of the cucumisin precursor. Cucumisin is the first known plant protease with such characteristics. Examination of the primary structure of cucumisin revealed that it is synthesized as a precursor, consisting of four functional domains: a possible signal peptide (22 amino acid residues), an NH2-terminal pro-sequence (88 residues), a 54-kDa protease domain (505 residues), which is the active enzyme domain of the 67-kDa native cucumisin, and a 14-kDa COOH-terminal polypeptide (116 residues), which arises by limited autolysis of the 67-kDa native cucumisin. This structure of cucumisin suggests that it is probably synthesized as an inactive precursor.
黄瓜蛋白酶是一种耐热的碱性丝氨酸蛋白酶，存在于甜瓜果实（甜瓜）的汁液中。我们已经确定了黄瓜蛋白酶cDNA的完整核苷酸序列（2552个核苷酸），并推导了相应的氨基酸序列。该cDNA的开放阅读框由731个密码子组成，编码一个相对于成熟天然黄瓜蛋白酶观察到的大小（67 kDa）较大的前体（分子量78815）。序列比较表明，黄瓜蛋白酶与枯草杆菌蛋白酶家族的微生物蛋白酶有几个共同特征。在黄瓜蛋白酶前体蛋白酶结构域的推导氨基酸序列中，可以发现与催化三联体氨基酸天冬氨酸、组氨酸和丝氨酸近端区域高度保守的序列，以及枯草杆菌蛋白酶中的底物结合位点。黄瓜蛋白酶是第一个已知具有这些特征的植物蛋白酶。对黄瓜蛋白酶一级结构的研究表明，它以前体形式合成，由四个功能结构域组成：一个可能的信号肽（22个氨基酸残基）、一个NH2末端前序列（88个残基）、一个54 kDa的蛋白酶结构域（505个残基），这是67 kDa天然黄瓜蛋白酶的活性酶结构域，以及一个14 kDa的COOH末端多肽（116个残基），它是由67 kDa天然黄瓜蛋白酶的有限自溶产生的。黄瓜蛋白酶的这种结构表明它可能以前体形式合成，是无活性的。
