ISOLATION OF A CHYMOTRYPSIN-LIKE ENZYME FROM TREPONEMA-DENTICOLA
ISOLATION OF A CHYMOTRYPSIN-LIKE ENZYME FROM TREPONEMA-DENTICOLA
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DOI：
10.1128/iai.56.10.2717-2722.1988
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发表时间：
1988-10-01
影响因子：
3.1
通讯作者：
MCBRIDE, BC
中科院分区：
文献类型：
--
作者：
UITTO, VJ;GRENIER, D;MCBRIDE, BC
从齿垢密螺旋体ATCC 35405中提取一种Mr为95，000的胰凝乳蛋白酶样蛋白酶，并通过制备型十二烷基硫酸钠-聚丙烯酰胺凝胶电泳进行部分纯化。蛋白水解活性在含有与牛血清白蛋白缀合的聚丙烯酰胺的层析图中检测。当T.在十二烷基硫酸钠的存在下溶解并溶解齿垢提取物。在文森特密螺旋体的提取物中没有发现活性。该酶水解转铁蛋白、纤维蛋白原、α 1-抗胰蛋白酶、免疫球蛋白A、免疫球蛋白G、明胶、牛血清白蛋白和含苯丙氨酸的合成肽。它不降解胶原蛋白或含有精氨酸或脯氨酸的合成底物。该酶水解偶氮唑的最适pH为7.5。在高于50 ℃的温度下加热。C破坏了活动。还原剂和螯合剂EDTA和乙二醇-双（β-氨基乙基醚）-N，N，N“，N”-四乙酸增加酶活性，而苯甲基磺酰氟、L-1-甲苯磺酰胺-2-苯乙基氯甲基酮、巯基试剂和人血清降低酶活性。该酶水解许多体液蛋白的能力表明它可能参与螺旋体的侵袭和组织破坏。
A chymotrypsinlike protease with an Mr of 95,000 was extracted from Treponema denticola ATCC 35405 and was partially purified by preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The proteolytic activity was detected in an electrophoretogram containing polyacrylamide that was conjugated to bovine serum albumin. A single band of activity was detected when the T. denticola extract was solubilized and electrophoresed in the presence of sodium dodecyl sulfate. No activity was found in extracts of Treponema vincentii. The enzyme hydrolyzed transferrin, fibrinogen, .alpha.1-antitrypsin, immunoglobulin A, immunoglobulin G, gelatin, bovine serum albumin, and a synthetic peptide containing phenylalanine. It did not degrade collagen or synthetic substrates containing arginine or proline. For the hydrolysis of azocoll, the pH optimum of the enzyme was 7.5. Heating at temperatures above 50.degree. C destroyed the activity. Reducing agents and the chelators EDTA and ethylene glycol-bis(.beta.-aminoethyl ether)-N,N,N'',N''-tetraacetic acid increased the enzyme activity, while phenylmethylsulfonyl fluoride, L-1-tosylamide-2-phenylethyl chloromethyl ketone, sulfhydryl reagents, and human serum reduced activity. The ability of the enzyme to hydrolyze a number of humoral proteins suggests that it may be involved in spirochete invasiveness and tissue destruction.