Birkedal-Hansen H, Taylor R E, Bhown A S, Katz J, Lin H Y, Wells B R
J Biol Chem. 1985 Dec 25;260(30):16411-7.
We have studied the susceptibility of fibrils formed from fetal bovine skin type III collagen to proteolytic enzymes known to cleave within the helical portion of the molecule (vertebrate and microbial collagenase, polymorphonuclear elastase, trypsin, thermolysin) and to two general proteases of broad specificity (plasmin, Pronase). Fibrils reconstituted from neutral salt solutions, at 35 degrees C, were highly resistant to nonspecific proteolysis by general proteases such as polymorphonuclear elastase, trypsin, and thermolysin but were rapidly dissolved by bacterial and vertebrate collagenases at rates of 12-45 mol X mol-1 X h-1. In solution, type III collagen was readily cleaved by each of the proteases (with the exception of plasmin), as well as by the true collagenases, although at different rates. Turnover numbers determined by viscometry at 35 degrees C were: human collagenase, approximately equal to 1500 h-1; microbial (clostridial) collagenase, approximately equal to 100 h-1; and general proteases, 23-52 h-1. In addition it was shown that pronase cleaves type III collagen in solution at 22 degrees C by attacking the same Arg-Gly bond in the alpha 1(III) chain as trypsin. However, like other proteases, Pronase was rather ineffective against fibrillar forms of type III collagen. It was also shown that transition of type III collagen as well as type I collagen to the fibrillar form resulted in a significant gain of triple helical thermostability as evidenced by a 6.8 degrees C increase in denaturation temperature (Tm = 40.2 degrees C in solution; Tm = 47.0 degrees C in fibrils).
我们研究了由胎牛皮肤III型胶原蛋白形成的原纤维对已知能在分子螺旋部分内切割的蛋白水解酶（脊椎动物和微生物胶原酶、多形核弹性蛋白酶、胰蛋白酶、嗜热菌蛋白酶）以及两种具有广泛特异性的通用蛋白酶（纤溶酶、链霉蛋白酶）的敏感性。在35℃下从中性盐溶液中重构的原纤维对多形核弹性蛋白酶、胰蛋白酶和嗜热菌蛋白酶等通用蛋白酶的非特异性蛋白水解具有高度抗性，但能被细菌和脊椎动物胶原酶以12 - 45 mol×mol⁻¹×h⁻¹的速率迅速溶解。在溶液中，III型胶原蛋白很容易被每种蛋白酶（纤溶酶除外）以及真正的胶原酶切割，尽管切割速率不同。在35℃下通过粘度测定法确定的转换数为：人胶原酶，约等于1500 h⁻¹；微生物（梭菌属）胶原酶，约等于100 h⁻¹；通用蛋白酶，23 - 52 h⁻¹。此外，研究表明链霉蛋白酶在22℃下通过攻击α1(III)链中与胰蛋白酶相同的精氨酸 - 甘氨酸键来切割溶液中的III型胶原蛋白。然而，与其他蛋白酶一样，链霉蛋白酶对III型胶原蛋白的纤维形式相当无效。研究还表明，III型胶原蛋白以及I型胶原蛋白向纤维形式的转变导致三螺旋热稳定性显著提高，变性温度升高6.8℃（溶液中Tm = 40.2℃；纤维中Tm = 47.0℃）证明了这一点。
