GELATION OF THE LENS CAPSULE TYPE-IV COLLAGEN SOLUTION AT A NEUTRAL PH

It is known that the type IV collagen extracted from EHS tumor assembl es under a physiological condition, but not in a gel form. The EHS typ e IV collagen requires the other basement membrane components, laminin 1, heparansulfate proteoglycan, and/or nidogen for gelation. On the ot her hand, Muraoka et al. reported that the bovine lens capsule type IV collagen alone gelated under a unique and unexpected condition of 2 M guanidine-HCl and 50 mM dithiothreitol, a condition which is thought to be dissociative for most biological macromolecules, including extra cellular matrix [Muraoka, M. et al. (1996) J. Biochem. 119, 167-172]. The present report shows that the bovine lens capsule type IV collagen formed a gel under physiological conditions of pH and ionic environme nt, though the apparent rigidity of the gel was weaker than that of th e gel formed in 2 M guanidine-HCl and dithiothreitol. The rigidity dep ended greatly on the incubation temperature and NaCl concentration of the type IV collagen solution, as observed in terms of the contractili ty of gel volume under centrifugal force. The gel formed in 150 mM NaC l and 20 mM phosphate, pH 7.3, at 28 degrees C contracted to 20% of th e original volume on centrifugation of 1,800 x g for 10 min, while the gel formed at 4 degrees C, where type I collagen did not gelate at al l, retained 90% of the original volume at the same centrifugal force. NaCl concentration was another important factor influencing the mechan ical properties of type IV collagen gel. The gel formed at 150 mM show ed maximal rigidity in the range of 0 to 300 mM in terms of the contra ctility on centrifugation. An image of a Pt/C replica of the gelated t ype IV collagen reconstituted at 4 or 28 degrees C in 20 mM phosphate, pH 7.3, containing 150 mM NaCl showed fine meshworks consisting of ra ther homogeneous pore sizes, resembling the skeletal structure of basa l lamina. Since the condition where the type IV collagen alone formed gels was physiological in terms of ionic strength and pH, the aggregat e structure and gel properties might reflect the in vivo type IV colla gen supramolecular structure and the property.