State Key Laboratory for Agrobiotechnology and Department of Microbiology, China Agricultural University, Beijing 100193, China.
J Biosci Bioeng. 2012 Jan;113(1):42-7. doi: 10.1016/j.jbiosc.2011.09.005. Epub 2011 Oct 20.
A novel serine protease, designated as cordysobin, was purified from dried fruiting bodies of the mushroom Cordyceps sobolifera. The isolation procedure utilized ion exchange chromatography on DEAE-cellulose and SP-Sepharose followed by gel filtration on Superdex 75. The protease did not adsorb on DEAE-cellulose but bound to SP-Sepharose. In sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), the protease resolved as a single band with an apparent molecular mass of 31 kDa. Its optimal pH was 10.0, and the optimal temperature was 65°C. The protease displayed a K(m) value of 0.41 μM and 13.44 μM·min⁻¹ using Suc-Leu-Leu-Val-Tyr-MCA as substrate at pH 10.0 and 37°C. Protease activity was enhanced by the Fe²⁺ ion at low concentration range of 1.25-10 mM and was strongly inhibited by Hg²⁺ up to 1.25 mM. The protease was strongly inhibited by chymostatin and phenylmethylsulfonyl fluoride (PMSF), suggesting that it is a serine protease. It manifested significant inhibitory activity toward HIV-1 reverse transcriptase (RT) with an IC₅₀ value of 8.2×10⁻³ μM, which is the highest anti-HIV-1 RT activity of reported mushroom proteins.
一种新型丝氨酸蛋白酶，命名为 cordysobin，从蛹虫草的干燥子实体中被分离出来。分离过程利用 DEAE-纤维素和 SP-Sepharose 离子交换层析，然后在 Superdex 75 上进行凝胶过滤。该蛋白酶不吸附在 DEAE-纤维素上，但与 SP-Sepharose 结合。在十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（SDS-PAGE）中，蛋白酶以 31 kDa 的表观分子量解析为单一带。其最适 pH 值为 10.0，最适温度为 65°C。该蛋白酶在 pH 10.0 和 37°C 下以 Suc-Leu-Leu-Val-Tyr-MCA 为底物时，K(m) 值分别为 0.41 μM 和 13.44 μM·min⁻¹。在低浓度范围（1.25-10 mM）内，Fe²⁺离子增强蛋白酶活性，Hg²⁺在 1.25 mM 时强烈抑制蛋白酶活性。该蛋白酶被糜蛋白酶和苯甲基磺酰氟（PMSF）强烈抑制，表明它是一种丝氨酸蛋白酶。它对 HIV-1 逆转录酶（RT）表现出显著的抑制活性，IC₅₀值为 8.2×10⁻³ μM，这是报道的蘑菇蛋白中抗 HIV-1 RT 活性最高的。
