National Resource for Automated Molecular Microscopy

DESCRIPTION (provided by the applicant): The National Resource for Automated Molecular Microscopy (NRAMM) was established in December 2002 to develop, test and apply technology for automating structure determination of macromolecules by cryoelectron microscopy (cryoEM). Since then, challenging collaborative projects have driven the development of several new techniques and technologies, which have subsequently been distributed to the scientific community. We demonstrated the feasibility of higher throughputs by acquiring ~300,000 single particles within 24 hours and established routine procedures for automated processing and analysis that, for well-behaved specimens, are capable of producing sub-nanometer resolution maps during the data acquisition session. With the basic infrastructure for automation now in place, our goals over the next five years are to add to and extend this infrastructure in order to provide a complete and integrated pipeline for the reconstruction of macromolecular machines. Our developments will provide us with a unique opportunity to systematically probe some of the current limitations of the methods and will result in more rational approaches to optimizing the quality and quantity of data acquisition and analysis. Core TRD projects are focused in 4 areas: improving grid substrates and specimen preparation;further automation and optimization of image acquisition;development of an integrated single particle analysis and processing pipeline that will be used to investigate parameters that limit resolution;and the development of automated high throughput EM screening. The four core projects will be driven by 13 collaborative projects;NRAMM will also continue to provide resources for about 30 service projects per year. Dissemination and Training activities will include the large International biennial NRAMM Practical Course in Molecular Microscopy as well as smaller training and multi-disciplinary workshops.