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| Content Provider | PubMed Central |
|---|---|
| Author | Kumaran, S. Datta, D. Roy, R. P. |
| Abstract | We have studied the conformation as well as V8 protease-mediated synthesis of peptide fragments, namely amino acid residues 295-316 (TC-peptide) of thermolysin and residues 1-20 (S-peptide) of ribonuclease A, to examine whether "conformational trapping" of the product can facilitate reverse proteolysis. The circular dichroism study showed cosolvent-mediated cooperative helix formation in TC-peptide with attainment of about 30-35% helicity in the presence of 40% 1-propanol and 2-propanol solutions at pH 6 and 4 degrees C. The thermal melting profiles of TC-peptide in the above cosolvents were very similar. V8 protease catalyzed the synthesis of TC-peptide from a 1:1 mixture of the non-interacting complementary fragments (TC295-302 and TC303-316) in the presence of the above cosolvents at pH 6 and 4 degrees C. In contrast, V8 protease did not catalyze the ligation of S1-9 and S10-20, although S-peptide could assume helical conformation in the presence of the cosolvent used for the semisynthetic reaction. V8 protease was able to synthesize an analog of S-peptide (SA-peptide) in which residues 10-14 were substituted (RQHMD-->VAAAK). While S-peptide exhibited helical conformation in the presence of aqueous propanol solutions, SA-peptide displayed predominantly beta-sheet conformation. SA-peptide showed enhanced resistance to proteolysis as compared with S-peptide. Thus, failure of semisynthesis of S-peptide may be a consequence of high flexibility around the 9-10 peptide bond due to its proximity to the helix stop signal. The results suggest that protease-mediated ligations may be achieved by design and manipulation of the conformational aspects of the product. |
| Starting Page | 2233 |
| File Format | |
| ISSN | 1469896X |
| e-ISSN | 1469896X |
| Journal | Protein Science : A Publication of the Protein Society |
| Issue Number | 10 |
| Volume Number | 6 |
| Language | English |
| Publisher | Cold Spring Harbor Laboratory Press |
| Publisher Date | 1997-10-01 |
| Access Restriction | Open |
| Rights Holder | Cold Spring Harbor Laboratory Press |
| Subject Keyword | Proteomics Research in Higher Education |
| Content Type | Text |
| Resource Type | Article |
| Subject | Medicine Molecular Biology Biochemistry |
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