March 2, Mon 2009
9:30 am, MRB 100 Conference Room
Complement 1s is the serine protease that cleaves IGFBP-5 in human osteoarthritic joint fluid
Insulin-like growth factor-I (IGF-I) and IGF binding proteins (IGFBPs) are trophic factors for cartilage and have been shown to be chondroprotective in animal models of osteoarthritis (OA). IGFBP-5 is degraded in joint fluid and inhibition of IGFBP-5 degradation has been shown to enhance the trophic effects of IGF-I. OBJECTIVE: To determine the identity of IGFBP-5 protease activity in human OA joint fluid. METHOD: OA joint fluid was purified and the purified material was analyzed by IGFBP-5 zymography. RESULTS: Both crude joint fluid and purified material contained a single band ofproteolytic activity that cleaved IGFBP-5. Immunoblotting of joint fluid for complement 1s (C1s) showed a band that had the same Mr estimate, e.g., 88kDa. In gel trypticdigestion and subsequent peptide analysis by LC-MS/MS showed that the band contained human C1s. A panel of protease inhibitors was tested for their ability to inhibit IGFBP-5 cleavage by the purified protease. Three serine protease inhibitors, FUT175 and CP-143217 and CB-349547 had IC50's between 1 and 6muM. Two other serine protease inhibitors had intermediate activity (e.g., IC50's 20-40muM) and MMP inhibitors had no detectible activity at concentrations up to 300muM. CONCLUSION: Human OA fluid contains a serine protease that cleaves IGFBP-5. Zymography, immunoblotting and LC-MS/MS analysis indicate that C1s is the protease that accounts for this activity.
10:00 am, MRB 100 Conference Room
Parul V. Kudtarkar
Roundup: Scaling computation to compute orthologs
Reciprocal smallest distance algorithm uses BLAST followed by evolutionary distance estimation based on pairwise global alignment to detect orthologs between genomes1. Round up is web application which uses RSD algorithm to find orthologs between query and subject genome(). Currently there are 220+ genomes in the database and 24,090 unique pairwise comparisons (i.e.n(n‐1)/2, where n is the number of genomes).Hence computation of orthologs is computationally intensive process, taking several CPU days. Cloud computing coupled with programming paradigm to parallelize computation of orthologs; significantly reduces the computation time and cost. The presentation will give overview of RoundUpand the parallelization of computation to calculate orthologs, which is my current research work at Center for Biomedical Informatics, Harvard Medical School.
10:30 am, MRB 100 Conference Room
INFLUENCE OF COOPERATIVITY ON THE PROTEIN FOLDING MECHANISM
Proteins fold reliably into a unique 3-dimensional structure essential to their biological function. My work aims to theoretically understand the mechanism of protein folding kinetics. I use a cooperative variational model and focus on two-state fast folding proteins. The predicted folding rates for 28 two-state fast folding proteins are correlated with experimental folding rates reasonable, and also the range of predicted folding rates is same as the wide range of measured folding rates which most models fail to predict correct range of folding rates since they have too low cooperativity to mimic real proteins. Moreover, the analysis of structure of transition state ensembles for cooperative and noncooperative cases shows that cooperativity sharpens the interface region between folded and unfolded region.
The cooperative variational model studied in this work is successful in predicting folding rates and structure of transition state ensembles for two state fast folding proteins.