The Collagen V Homotrimer Production Is Unexpectedly Favored over the Heterotrimer in Recombinant Expression Systems
Figure 6
(a) Selection of clones expressing pro1(V), pro2(V) chains and HSP47. Western blot analysis of transfected cell media (for pro1(V) and pro2(V) chain detection) or cell lysates (for HSP47 detection) probed with (first panel) 6A7 monoclonal antibody, which recognizes the N-propeptide of the human pro1(V) chain, (second panel) with collagen type V (G-15) antibodies, which recognize the human pro2(V) chain, (third panel) with antibody to HSP47. Of twenty-one clones, only five clones expressed the three partners with different expression levels: C2 (lane 1), C7 (lane 2), C9 (lane3), C13 (lane 4), and C15 (lane 5). The other clones expressed only two of the three partners. For example, clone C12 (lane 6) expressed only the pro1(V) chain and HSP47. (b) Western blot analysis of media from clone C2 treated with (lanes 3, 4) or without monensin (lanes 1, 2) using polyclonal collagen V antibodies. Media were incubated with (lanes 2, 4) or without pepsin (lanes 1, 3). Left, molecular mass standards expressed in kDa.
