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INVESTIGATION OF DYNEIN’S LINKER REGIONSMentor: Dr. Elisar Barbar
Jonathan Yih
RELEVANCE
Role in basic life processes
http://hopes.stanford.edu/basics/dna/f_b14mit_meta.jpg
Insight for dynein-associated problems
http://www.thehealthsuccesssite.com/images/muscle-als.jpg
GOAL
Improve understanding of dynein’s structure
http://www.physics.berkeley.edu/research/yildiz/DyneinNew.jpg
BACKGROUND
Motor protein
Energy Work
Functions:
Transport
Orienting & positioning organelles
Cell division http://maepro.uta.edu/loco/pictures/kinesin11.jpg
BACKGROUND
Dynein’s structure
• Heavy Heads & Chains
ATP hydrolysis & movement
• Light & Intermediate Chains
Binding
IC
LC8
Cargo
Heavy Heads and Chains
Mic
rotu
bu
le
HYPOTHESIS
LC8 binding to IC initiates the assembly of the dynein complex.
OVERVIEW
Surface Plasmon ResonanceStudy protein kinetics
LC8 with:IC
nNOS
BIM
Swa
LC8/IC compex image © Barbar 2007
SURFACE PLASMON RESONANCE
Gold Surface
Results!
Immobilized Protein (LC8)
Incident Light Reflected Light
Analyte
SURFACE PLASMON RESONANCE
http://www.discoversensiq.com/uploads/file/support/spr/Overview_of_SPR.pdf
Protein complex formation
A + B ABka
kd
SURFACE PLASMON RESONANCE
Immobilization
1
2
3
Time (s)
SPR: LC8 & BIM OR NNOS
SPR: LC8 & IC
Res SD 6.704
SPR: EXPERIMENTAL DESIGN
Replications
Multiple concentrations (5+)
Adequate regeneration
Reduce mass transport limitations
Achieve steady state
ACKNOWLEDGEMENTS
Special thanks to:Dr. Elisar BarbarThe Barbar Lab Group
Afua Nyarko, Yujuan Song, & Jessica Morgan
Nathan Gillock & Aaron MartinDr. Kevin AhernThe Howard Hughes Medical Institute