Moss J M, Van Damme M P, Murphy W H, Stanton P G, Thomas P, Preston B N
Department of Biochemistry and Molecular Biology, Monash University, Clayton, Victoria, Australia.
Arch Biochem Biophys. 1997 Mar 1;339(1):172-82. doi: 10.1006/abbi.1996.9828.
The cationic protein, lysozyme, has an extracellular distribution in cartilage; however, its biological role in this tissue still remains unclear. This study describes a simple and high yielding procedure for the purification of four novel isoforms of lysozyme from the functionally different articular (metacarpalphalangeal joint) and nonarticular (nasal septum) bovine cartilages. Chromatography of the cartilage extracts on S-Sepharose revealed the presence of four major lysozyme active peaks each of which was further purified to homogeneity by gel filtration and reversed-phase chromatography. Each peak yielded a different molecular mass when analyzed by ion spray mass spectrometry, and material isolated from either cartilage source displayed an identical molecular mass for each lysozyme preparation. N-terminal amino acid sequence and amino acid composition analyses confirmed the presence of four novel lysozyme isoforms in both bovine articular and nonarticular cartilages. The lytic activity of each lysozyme isoform toward Micrococcus lysodeikticus was dependent on both the ionic strength and pH of the buffer, where an increase in activity accompanied an increase in ionic strength. The lysozymes were shown to be synthesized by chondrocytes in vitro, which in addition to the relatively high chemical amounts of lysozyme present in cartilage, would suggest that this small cationic protein has some as yet undetermined biological role within the cartilage extracellular matrix.
阳离子蛋白溶菌酶在软骨中呈细胞外分布；然而，其在该组织中的生物学作用仍不清楚。本研究描述了一种简单且高产的方法，用于从功能不同的牛关节（掌指关节）和非关节（鼻中隔）软骨中纯化四种新型溶菌酶同工型。软骨提取物在S-Sepharose上进行色谱分析，显示出四个主要的溶菌酶活性峰，每个峰通过凝胶过滤和反相色谱进一步纯化至均一。通过离子喷雾质谱分析时，每个峰产生不同的分子量，并且从任一软骨来源分离的物质对于每种溶菌酶制剂显示相同的分子量。N端氨基酸序列和氨基酸组成分析证实了牛关节软骨和非关节软骨中均存在四种新型溶菌酶同工型。每种溶菌酶同工型对溶壁微球菌的裂解活性取决于缓冲液的离子强度和pH值，活性的增加伴随着离子强度的增加。溶菌酶在体外由软骨细胞合成，这除了软骨中存在相对较高化学量的溶菌酶外，表明这种小的阳离子蛋白在软骨细胞外基质中具有一些尚未确定的生物学作用。
