Human type V collagen was purified from placenta and found to contain α1(V), α2(V), and α3(V) chains in varying ratios. Using any of three independent non-denaturing methods (phosphocellulose chromatography, high-performance ion-exchange chromatography on IEX-540 DEAE, and ammonium sulfate precipitation), this preparation could be resolved into two fractions. Analysis of the two fractions by sodium dodecyl sulfate-polyacrylamide gel electrophoresis indicated that one fraction contained α1(V) and α2(V) in a 2:1 ratio and the other contained α1(V), α2(V), and α3(V) in a 1:1:1 ratio. When the crude placental type V collagen was electrophoresed under nondenaturing conditions, two bands were observed, one co-migrating with purified (α1(V))2α2(V) and the other co-migrating with the fractions containing α1(V), α2(V), and α3(V) chains in a 1:1:1 ratio. Electrophoresis in a second dimension under denaturing conditions confirmed that the fast-migrating band contained (α1(V))2α2(V) and that the slow-migrating band contained the three chains in equimolar ratio. CD spectra of the two fractions and resistance to trypsin-chymotrypsin digestion confirmed that the two fractions contain triple helical collagen. Thermal denaturatons were monitored by the changes in CD signal at 221 nm. The two fractions purified by ammonium sulfate precipitation melted at 39.1 and 36.4° C for the (α1(V)2α2(V) and α1(V)α2(V)α3(V) fractions, respectively. Trypsin cleavage of these two native fractions at temperatures near melting produced completely different fragmentation patterns, indicating different partial unwinding sites of the α1(V) and α2(V) chains in the two preparations and thus different molecular assemblies. Our data demonstrate the existence of two different molecular assemblies of type V collagen in human placenta consisting of (α1(V))2α2(V) and α1(V)α2(V)α3(V) heterotrimers.
|Original language||English (US)|
|Number of pages||5|
|Journal||Journal of Biological Chemistry|
|State||Published - Dec 1 1984|
All Science Journal Classification (ASJC) codes
- Molecular Biology
- Cell Biology