Purification, cloning, and expression of human plasma hyaluronidase
Purification, cloning, and expression of human plasma hyaluronidase
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DOI：
10.1006/bbrc.1997.6773
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发表时间：
1997-07-09
影响因子：
3.1
通讯作者：
Stern, R
中科院分区：
文献类型：
--
作者：
Frost, GI;Csoka, TB;Stern, R
使用 Triton X-114 相萃取和离子交换色谱从人血浆中纯化透明质酸酶。通过一种新颖的筛选测定法产生针对纯化蛋白的单克隆抗体，仅用于分离均质酶恐惧微测序，获得的氨基酸序列与表达序列标签数据库中的cDNA相匹配，该数据库通过5'-RACE-PCR单独用于血浆透明质酸酶基因，称为Hgal-1。 Hyal-1 编码由 435 个氨基酸组成的蛋白质，与精子特异性透明质酸酶 PH-20 的同一性超过 40%。与仅在睾丸中表达的 PH-20 不同，Hyal-1 的转录本在多个组织中都有发现，Hyal-1 在人胚胎肾细胞中稳定表达，导致分泌的免疫反应性透明质酸酶活性增加 3,000 倍，这在生化上与人血浆透明质酸酶无法区分。通过免疫学、分子和生化标准，我得出结论，Hyal-1 是主要的人类血浆中发现的透明质酸酶。 (C) 1997 年学术出版社。
Hyaluronidase was purified from human plasma using Triton X-114 phase extractions and ion-exchange chromatography. Monoclonal antibodies generated against the purified protein by a novel screening assay mere utilized to isolate homogeneous enzyme fear microsequencing, The amino acid sequences obtained matched a cDNA in the Expressed Sequence Tag database which, with 5'-RACE-PCR, was used to alone the plasma hyaluronidase gene, termed Hgal-1. Hyal-1 codes for a protein of 435 amino acids that is over 40% identical to PH-20, sperm-specific hyaluronidase. Unlike PH-20, which is only expressed in testis, transcripts of Hyal-1 were found in multiple tissues, Hyal-1 stably expressed in human embryonic kidney cells resulted in a 3,000 fold increase of secreted immunoreactive hyaluronidase activity that was biochemically indistinguishable from human plasma hyaluronidase, By immunological, molecular and biochemical criteria, me conclude that Hyal-1 is the predominant hyaluronidase found in human plasma. (C) 1997 Academic Press.