Figure 1.

Elastase and SMART^TM methods led to the improved detection of fibrillin-1 and improved identification of microfibril-associated proteins compared with previous published efforts. The ability of the elastase method to produce peptides of fibrillin-1 from a single human CB sample (female age 67; F67) and a single human skin sample (F55) is compared with the efforts of Cain et al. (23) (A). As performed by Cain et al. (23), PSMs (Peptide Prophet FDR ≤ 5%) were counted for each respective fibrillin-1 domain and heat-mapped. Our method led to a greater primary coverage and domain coverage from a single sample run than Cain et al. (23), whose coverages were achieved from 13 separate sample runs. Peptides from the C-terminal region of fibrillin-1 were also successfully detected (orange arrow), which Cain et al. (23) failed to identify. The known fibrillin microfibril-interacting proteins identified by Cain et al. (23) (green) are compared with those identified by elastase and SMART^TM methods (Protein Prophet FDR ≤ 0.1%, Peptide Prophet FDR ≤ 5%) (B). The elastase method (red) and SMART^TM method (blue) were both performed on the same human CB microfibril extract (F73). The elastase method appears to enhance the detection of microfibril-associated proteins thought to be tightly bonded to the structure (i.e. the MFAP family), whereas the SMART^TM method appears to enhance the detection of weakly interacting proteins (i.e. versican and hyaluronan proteins). Collectively, these methods led to an enhanced detection of known microfibril-associated proteins compared with Cain et al. (23).