Transient collagen triple-helix binding to membrane type 1 matrix metalloproteinase : interaction studies and NMR-guided structural docking

[ACCESS RESTRICTED TO THE UNIVERSITY OF MISSOURI AT AUTHOR'S REQUEST.] MT1-MMP (MMP-14) as pericellular collagenase is critically involved in cancer cell invasion through collagen barriers that it degrades. To better understand the structural and mechanistic details underlying collagenolytic activity of MMP-14, a solution NMR approach is used here to investigate interactions between individual MMP-14 catalytic and hemopexin domain and a collagen-I-mimicking Triple-Helical Peptide (THP). In this study, backbone chemical shifts were assigned for isolated MMP-14 catalytic and hemopexin domains. The results from gel-filtration chromatography, DLS and NMR combined suggested that MMP-14 hemopexin domain behaves consistently as a monomer in solution. NMR-monitored THP titration led to identification of a distinct patch centered about blade I at the exit side of the hemopexin domain as a potential THP binding exosite. Mutation of residues from this area impairs triple-helical peptidase activity of MMP-14. Saturation transfer difference NMR suggests rotational averaging around the longitudinal axis of the triple-helical peptide. Additionally, intermolecular distances between the hemopexin domain of MMP-14 (HPX-14) and THP were measured by paramagnetic NMR using TOAC-labeled THP. Structural models of HPX-14/THP calculated based on PRE-measured distance restraints revealed extensive interaction between MMP-14 hemopexin domain and sequences surrounding the cleavage site in the THP, indicating a distinctive arrangement of the catalytic domain and unique collagen binding conformation of MMP-14 during collagenolysis.

Name thesis:degree_name

Ph. D.

Level thesis:degree_level

Doctoral

Discipline thesis:degree_discipline

Biochemistry (MU)

Grantor dc:publisher

University of Missouri--Columbia

Year dc:date.issued

2015

Author dc:creator

• Zhao, Yingchu

Advisor dc:contributor.advisor

• Van Doren, Steven R., 1963-

Statement dc:rights

• Access to files is limited to the campuses of the University of Missouri with SSO login.

Language dc:language.iso

eng, English

DOI

https://doi.org/10.32469/10355/49038

OAI identifier oai:identifier

oai:mospace.umsystem.edu:10355/49038