Figure. The binding activity of COL6a1 with RTBDN.
The collagens are a superfamily of proteins that play a role in maintaining the integrity of various tissues. Collagen VI is a major structural component of microfibrils. The basic structural unit of collagen VI is a heterotrimer of the alpha1(VI), alpha2(VI), and alpha3(VI) chains. The protein encoded by COL6a1 gene is the alpha 1 subunit of type VI collagen (alpha1(VI) chain). Mutations in the genes that code for the collagen VI subunits result in the autosomal dominant disorder, Bethlem myopathy. Besides, Retbindin (RTBDN) has been identified as an interactor of COL6a1, thus a binding ELISA assay was conducted to detect the interaction of recombinant mouse COL6a1 and recombinant mouse RTBDN. Briefly, COL6a1 were diluted serially in PBS, with 0.01% BSA (pH 7.4). Duplicate samples of 100uL were then transferred to RTBDN-coated microtiter wells and incubated for 2h at 37℃. Wells were washed with PBST and incubated for 1h with anti-COL6a1 pAb, then aspirated and washed 3 times. After incubation with HRP labelled secondary antibody, wells were aspirated and washed 3 times. With the addition of substrate solution, wells were incubated 15-25 minutes at 37℃. Finally, add 50μL stop solution to the wells and read at 450nm immediately. The binding activity of COL6a1 and RTBDN was shown in Figure 1, and this effect was in a dose dependent manner.