Use of transmission electron microscopy to identify nanocrystals of challenging protein targets

Hilary P. Stevenson; Alexander M. Makhov; Monica CaleroAndrea L. EdwardsOliver B. ZeldinIrimpan I. MathewsGuowu LinChristopher O. BarnesHugo SantamariaTed M. RossS. Michael SoltisChaitan KhoslaV. NagarajanJames F. ConwayAina E. CohenGuillermo Calero

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Use of transmission electron microscopy to identify nanocrystals of challenging protein targets

机译：Use of transmission electron microscopy to identify nanocrystals of challenging protein targets

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The current practice for identifying crystal hits for X-ray crystallography relies on optical microscopy techniques that are limited to detecting crystals no smaller than 5 μm. Because of these limitations, nanometer-sized protein crystals cannot be distinguished from common amorphous precipitates, and therefore go unnoticed during screening. These crystalswould be ideal candidates for further optimization or for femtosecond X-ray protein nanocrystallography. The latter technique offers the possibility to solve high-resolution structures using submicron crystals. Transmission electron microscopy (TEM) was used to visualize nanocrystals (NCs) found in crystallization drops that would classically not be considered as "hits." We found that protein NCs were readily detected in all samples tested, includingmultiprotein complexes andmembrane proteins. NC quality was evaluated by TEM visualization of lattices, and diffraction quality was validated by experiments in an X-ray free electron laser.

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《Proceedings of the National Academy of Sciences of the United States of America》 |2014年第23期|8470-8475|共6页
作者
Hilary P. Stevenson; Alexander M. Makhov; Monica CaleroAndrea L. EdwardsOliver B. ZeldinIrimpan I. MathewsGuowu LinChristopher O. BarnesHugo SantamariaTed M. RossS. Michael SoltisChaitan KhoslaV. NagarajanJames F. ConwayAina E. CohenGuillermo Calero;
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作者单位

Departments of Chemical Engineering and Chemistry, Stanford University, Stanford, CA 94305;

Department of Structural Biology, Stanford University, Stanford, CA 94305;

Vaccine and Gene Therapy Institute of Florida, Port St. Lucie, FL 34987Stanford Synchrotron Radiation Lightsource, SLAC National Accelerator Laboratory, Stanford University, Menlo Park, CA 94025Department of Structural Biology, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261JAN Scientific, Inc., Seattle, WA 98105;

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收录信息美国《科学引文索引》(SCI);美国《生物学医学文摘》(MEDLINE);美国《化学文摘》(CA);
原文格式 PDF
正文语种英语
中图分类自然科学总论;
关键词
nanocrystal detection; crystal optimization; crystal characterization; femtosecond diffraction; structural biology;

Use of transmission electron microscopy to identify nanocrystals of challenging protein targets

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