Dean M F, Sansom P
Department of Veterinary Basic Sciences, The Royal Veterinary College, University of London, Royal College Street, London NW1 0TU, UK.
Biochem J. 2000 Jul 15;349(Pt 2):473-9. doi: 10.1042/0264-6021:3490473.
The peptide DHLSDNYTLDHDRAIH (Link N), cleaved from the N-terminus of the link protein component of cartilage proteoglycan aggregates by the action of stromelysin, can act as a growth factor and stimulate synthesis of proteoglycans and collagen in articular cartilage [McKenna, Liu, Sansom and Dean (1998) Arthritis Rheum. 41, 157-161]. The mechanism by which this biologically active peptide is degraded and inactivated was investigated using U937 monocytes as a model cell. Time-course experiments showed that two major proteases, an initial serine proteinase followed by a metalloproteinase, acted in sequence. Analysis of the resulting fragments showed that the serine endopeptidase cleavage was at the Leu(3)-Ser(4) bond to produce the peptide SDNYTLDHDRAIH. The terminal serine could then be removed from the resulting peptide by an aminopeptidase. A second metallopeptidase liberated the peptides SDNYTL or DNYTL from DHDRAIH by cleavage at the Leu(9)-Asp(10) bond. The DNYTL peptide intermediate was degraded too rapidly to allow sequencing and sequential aminopeptidase cleavages removed further amino acids from the N-terminus of the remaining DHDRAIH peptide. The identical patterns of breakdown that occurred when either whole cells or purified plasma membranes were used indicated that proteolysis and inactivation of Link N was carried out entirely by membrane-associated enzymes.
肽DHLSDNYTLDHDRAIH（连接N），在基质溶素的作用下从软骨蛋白聚糖聚集体的连接蛋白成分的N端裂解而来，可作为一种生长因子，刺激关节软骨中蛋白聚糖和胶原蛋白的合成[麦肯纳、刘、桑瑟姆和迪恩（1998年）《关节炎与风湿病》41卷，157 - 161页]。以U937单核细胞作为模型细胞，研究了这种生物活性肽被降解和失活的机制。时间进程实验表明，两种主要的蛋白酶依次起作用，首先是一种丝氨酸蛋白酶，随后是一种金属蛋白酶。对产生的片段进行分析表明，丝氨酸内肽酶的切割发生在Leu(3)-Ser(4)键处，产生肽SDNYTLDHDRAIH。然后，一种氨肽酶可以从产生的肽中去除末端丝氨酸。第二种金属肽酶通过在Leu(9)-Asp(10)键处切割，从DHDRAIH中释放出肽SDNYTL或DNYTL。DNYTL肽中间体降解太快，无法进行测序，并且连续的氨肽酶切割从剩余的DHDRAIH肽的N端去除了更多氨基酸。使用全细胞或纯化的质膜时出现的相同降解模式表明，连接N的蛋白水解和失活完全由膜相关酶进行。
