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【目的】桑氏链霉菌(Streptomyces sampsonii)KJ07对无性阶段的杨树紫纹羽病菌(Rhizoctonia violacea)有较强的拮抗作用。为研究其抗菌物质,对其发酵液中主要抗菌物质进行分离纯化并明确其部分性质。【方法】采用硫酸铵分级沉淀、DEAE Sepharose Fast Flow离子交换层析、Sephadex G-50分子筛层析等方法进行分离纯化。【结果】获得单一抗菌活性蛋白,分子量约为28.4 k D。该抗菌蛋白抑菌谱较广,能使R.violacea菌丝畸形,菌丝隔膜不明显,细胞壁及胞内原生质开始降解,并产生黑色物质。稳定性试验显示抗菌蛋白最适温度为25°C,最佳p H为6.0,当温度≥60°C时,抑菌活性下降大于20%,当p H<4.0或≥8.0时,抑菌活性下降大于12%。其活性还受金属阳离子影响,但对蛋白酶K不敏感。利用自动Edman降解法测得抗菌蛋白N端10个氨基酸序列,但通过NCBI BLAST程序未检索到与其相似性较高的已知抗菌蛋白。【结论】推测该抗菌蛋白可能是一种新的蛋白质。
【Objective】 Streptomyces sampsonii KJ07 has strong antagonism against Rhizoctonia violacea during the vegetative stage. In order to study its antibacterial substances, the main antibacterial substances in its fermentation broth were separated and purified and their partial properties were clarified. 【Method】 Separation and purification were carried out by ammonium sulfate fractionation, DEAE Sepharose Fast Flow ion exchange chromatography and Sephadex G-50 molecular sieve chromatography. 【Result】 Single antimicrobial active protein was obtained with molecular weight of about 28.4 kD. The antimicrobial protein antibacterial spectrum broader, can deformity R.violacea mycelium, hyphae membrane is not obvious, cell wall and intracellular protoplasts began to degrade, and produce black substances. The stability test showed that the optimum temperature of antimicrobial protein was 25 ° C and the optimal pH was 6.0. When the temperature was above 60 ° C, the antibacterial activity decreased by more than 20%. When p H <4.0 or ≥8.0, the antibacterial activity Fall more than 12%. Its activity is also affected by metal cations, but it is not sensitive to proteinase K. The N-terminal 10-amino acid sequence of the antimicrobial protein was detected by the automatic Edman degradation method. However, the known antimicrobial protein with high similarity was not found by the NCBI BLAST program. 【Conclusion】 It is speculated that the antimicrobial protein may be a new protein.