Troponin C - the calcium switch in muscle: Interactions
of TnC with TnI and their role in thin filament regulation
The Faculty of the Graduate School of Arts and Sciences
Professor Carolyn Cohen, Advisor
In Partial Fulfillment
of the Requirements for the Degree
Doctor of Philosophy
Michael L. Love
Troponin C - the calcium switch in muscle:
Interactions of TnC with TnI and their role in
thin filament regulation
and cardiac muscles are regulated by the tropomyosin/troponin complex.
Troponin C (TnC), a subunit of troponin, is the Ca2+ receptor
that controls regulation. The structure of recombinant skeletal TnC was
determined by x-ray crystallography in the four calcium bound "on state"
to 2.0 Å resolution. A comparison of two and four calcium bound
TnC structures reveals the interactions that must change during the Ca2+-dependent
transition. Sequence variations are shown to account for functional
differences in the calmodulin superfamily. In addition, we have solved
to 2.0 Å resolution the crystal structure of skeletal TnC complexed
with a synthetic peptide which comprises TnI residues Asn96-Lys123. This
sequence includes the inhibitory region, TnI(Gly104-Arg115), which can
also bind to actin. The structure reveals the interaction of residues
Leu111-Asp119 of TnI with TnC and may represent key structural characteristics
of the troponin "on state". A comparison of the structures of peptide
bound skeletal TnC and a drug bound cardiac TnC indicates that the peptide
and drug bind to different surfaces of TnC. This information may guide
the design of new drugs in the treatment of congestive heart failure and
other myopathic diseases.
List of Tables
Many thanks go to my fellow lab members, who helped me in science, in writing, in conversation, and so on. Thank you Yu Li, Jerry Brown, Suet Mui. Special thanks go to Anne Houdusse, who brought her exceptional talents of structural analysis to the problems we faced in Chapter 2, and to Roberto Dominguez, who taught me the practical protein crystallographic method and much more.
Also, thanks go to the Brandeis detector group. In particular, thank you Walter Phillips, Marty Stanton and Charlie Ingersoll, who taught me about the hardware and software of crystallographic data collection. I solved my first structure using data from an x-ray camera that we assembled.
I am very grateful to Zenon Grabarek, our collaborator, who not only provided the protein and peptides that were used in these experiments, but also provided many biochemical insights into the problems we were facing. I am also very grateful Carolyn Cohen, my advisor and teacher, who made all of this possible and gave me a chance to become a scientist. Many Thanks!