Center for Genomic Experimentation and Computation Orna Resnekov Molecular Sciences Institute Berkeley, CA

Magritte Goal: To combine genomic and computational research in order to make predictive models of biological systems. molecular sciences institute

a  a factor  factor zygote a/  The mating pathway

molecular sciences institute a  a factor  factor zygote a/  The Alpha Project

molecular sciences institute Pathway after pheromone We are trying to both model and simulate the pathway

molecular sciences institute Simplified view of pathway

molecular sciences institute  Abundance (Ste5 ~500, Fus3 ~20,000)  Reaction rates  Affinities of proteins for one another  Structural information (not covered in our current Center grant) To simulate we need numbers…

molecular sciences institute Key questions  What is the oligomerization state(s) of purified active Ste5?  What is the stoichiometry of the different multiprotein complexes that have kinase activity?  Which of the structures are biologically relevant Ste5 complexes?

molecular sciences institute Complexity of the sample and purification status  Ste5, Ste11, Ste7, Fus3  We have expression constructs to make individual proteins and reconstitute the pathway  We are also purifying complexes from cells using affinity tagged proteins (TAP) We have polyclonal abs, we are identifying posttranslational modifications (MS), we are making monoclonal abs to modified proteins, we have fluorescently tagged proteins.

molecular sciences institute Hurdles to overcome  Need a good partner for structural studies  We would like to be doing both cryo-EM and crystallography  Sufficient material for these studies

molecular sciences institute Resources needed  RO1 level funding for 3-5 people

molecular sciences institute Response to pheromone

molecular sciences institute The Team: Kirsten Benjamin Roger Brent-DIRECTOR Ian Burbulis Tina Chin Alejandro Colman-Lerner Andrew Gordon (P) Larry Lok (Math) Joyce Maccabea Gustavo Pesce David Pincus Orna Resnekov Dan Rockwell Eduard Serra David Soergel (CS) Kouichi Takahashi Kumiko Yamaguchi Richard Yu (Struct. Bio.) Myron Williams PNNL Richard Smith (Chem) Robert Maxwell SBML Andrew Finney U. WA. Dee Dee Meldrum Mark Holl MIT Drew Endy (ex MSI) Ty Thomson (BioEng) Caltech Shuki Bruck (CS) Mark Reidel Matt Cook