Assembly of E. coli ribosomes in vitro and in cells

Scripps Research Institute, The, La Jolla CA

Investigators

Linked publications & trials

Abstract

Principal Investigator/Program Director (Last, first, middle): Williamson, James, R. RESEARCH &RELATED Other Project Information 1. * Are Human Subjects Involved? m Yes l No 1.a. If YES to Human Subjects Is the IRB review Pending? m Yes m No IRB Approval Date: Exemption Number: 1 2 3 4 5 6 Human Subject Assurance Number 2. * Are Vertebrate Animals Used? m Yes l No 2.a. If YES to Vertebrate Animals Is the IACUC review Pending? m Yes m No IACUC Approval Date: Animal Welfare Assurance Number 3. * Is proprietary/privileged information m Yes l No included in the application? 4.a.* Does this project have an actual or potential impact on m Yes l No the environment? 4.b. If yes, please explain: 4.c. If this project has an actual or potential impact on the environment, has an exemption been authorized or an environmental assessment (EA) or environmental impact statement (EIS) been performed? m Yes m No 4.d. If yes, please explain: 5.a.* Does this project involve activities outside the U.S. or m Yes l No partnership with International Collaborators? 5.b. If yes, identify countries: 5.c. Optional Explanation: 6. * Project Summary/Abstract 5035-Project_Description.pdf Mime Type: application/pdf 7. * Project Narrative 3432-Project_Narrative.pdf Mime Type: application/pdf 8. Bibliography &References Cited 2163-literature.pdf Mime Type: application/pdf 9. Facilities &Other Resources 6727-Fac_&_Resources.pdf Mime Type: application/pdf 10. Equipment 8190-Equipment.pdf Mime Type: application/pdf Tracking Number: Other Information Page 5 OMB Number: 4040-0001 Expiration Date: 04/30/2008 Principal Investigator/Program Director (Last, first, middle): Williamson, James, R. Project Description This proposal outlines a plan to investigate the mechanism of assembly of E. coli ribosomes both in vitro and in vivo. The ribosome is the macromolecular machine responsible for all protein synthesis in cells, and it is efficiently assembled from over 50 components. Using a combination of biophysical methods, we will investigated the order of events in assembly and the kinetics of assembly in vitro. We will examine the role of assembly cofactors in E. coli, and examine the effects of antibiotics on the assembly process. We have developed a novel isotope pulse-chase assay that enables measurement of binding kinetics for ribosomal proteins using mass spectrometry. We are developing a two-photon fluorescence correlation microscope to monitor the assembly of fluorescently labeled ribosomal proteins in real time. Finally, we are extending our mass spectrometry analysis to isotope pulse experiments in living bacteria, which allows us to directly monitor the biogenesis of ribosomes. Taken together, these studies will provide new mechanistic insights into the critical process of ribosome assembly. Project Description Page 6