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Cambridge Institute for Medical Research

 

Methods for structural biology

General audience summary:
The three-dimensional structure of a protein can be determined using two techniques termed X ray crystallography and cryo-electron microscopy. This information provides vital insights into how a protein functions and how it is controlled by other factors in the cell. Our goal is to develop new improvements to these techniques, including the use of new software and computer modelling approaches, to allow more challenging structures to be elucidated. The techniques we are developing contribute to understanding the structures of proteins that affect disease development.

Strategic CIMR theme: Protein folding and quality control

Funding: Wellcome Trust, National Institutes of Health (USA)

Research Group members: Tristan Croll, Airlie McCoy, Claudia Millán, Robert Oeffner, Duncan Stockwell

Biography

Randy Read was born in Canada and studied Biochemistry at the University of Alberta in Edmonton, specialising in protein crystallography for his PhD. After a post-doctoral fellowship at the University of Groningen in The Netherlands, he took up his first faculty position in the Department of Medical Microbiology and Immunology, supported by the Medical Research Council of Canada and the Howard Hughes Medical Institute.  In 1998 he moved to Cambridge to join the new Cambridge Institute for Medical Research, supported by a Wellcome Trust Principal Research Fellowship.

Research

Methods for structural biology

Developing improved methods for determining structures of proteins and other macromolecules. 

For decades, crystallography has been the primary method for determining the three-dimensional structure of a protein, which provides an essential framework for a detailed understanding of its biochemistry. We have been focusing on extending the scope and power of the methods used in protein crystallography. With the recent revolution in the ability of cryo-electron microscopy methods to determine structure at high resolution, we are now also looking at the intersection between these techniques.

In crystallographic theory, we focus on the understanding of probability distributions relating the structure factors that arise from a diffraction experiment. A detailed understanding of these probability distributions underlies new developments in maximum likelihood methods, which we are implementing in our program Phaser. The current version of Phaser can solve structures by molecular replacement (that is, using the known structures of related proteins), by using the information from single-wavelength anomalous diffraction (SAD), and by a combination of the two. By accounting better for the effects of errors, these new methods can cope with trends to collect data from poorer crystals to higher resolution, and can solve structures that evaded earlier approaches.

Cryo-electron microscopy shares some of the theoretical underpinnings of crystallography, since both methods involve analyses in Fourier space. As a result, we can now explore ways in which the lessons we learned in implementing likelihood methods for crystallography can be transposed to the new methods in cryo-EM.

Publications

Key publications: 

Methods

Terwilliger TC, Ludtke SJ, Read RJ, Adams PD and Afonine PV. Improvement of cryo-EM maps by density modification. Nature Methods 17, 923-927 (2020).

Read RJ, Oeffner RD and McCoy AJ. Measuring and using information gained by observing diffraction data. Acta Cryst. D76, 238-247 (2020).

Afonine PV, Boon BK, Read RJ, Sobolev OV, Terwilliger TC, Urzhumtsev A & Adams PD. Real-space refinement in PHENIX for cryo-EM and crystallography. Acta Cryst. D74, 531-544 (2018).

Oeffner RD, Afonine PV, Millán C, Sammito M, Usón I, Read RJ & McCoy AJ. On the application of the expected log-likelihood gain to decision making in molecular replacement. Acta Cryst. D74, 245-255 (2018).

McCoy AJ, Oeffner RD, Wrobel AG, Ojala JR, Tryggvason K, Lohkamp B & Read RJ. Ab initio solution of macromolecular crystal structures without direct methods. Proc. Natl. Acad. Sci. USA 114, 3637-3641 (2017).

Read RJ & McCoy AJ. A log-likelihood-gain intensity target for crystallographic phasing that accounts for experimental error. Acta Cryst. D72, 375-387 (2016).

Bunkóczi G, McCoy AJ, Echols N, Grosse-Kunstleve RW, Adams PD, Holton JM, Read RJ & Terwilliger TC. Macromolecular X-ray structure determination using weak, single-wavelength anomalous data. Nature Methods 12, 127–130 (2015).

Jackson RN, McCoy AJ, Terwilliger TC, Read RJ & Wiedenheft B. X-ray structure determination using low-resolution electron microscopy maps for molecular replacement. Nature Protocols 10, 1275-1284 (2015).

DiMaio F, Terwilliger TC, Read R., Wlodawer A, Oberdorfer G, Wagner U, Valkov E, Alon A, Fass D, Axelrod HL, Das D, Vorobiev SM, Iwaï H, Pokkuluri PR & Baker D. Improving molecular replacement by density- and energy-guided protein structure optimization. Nature 473, 540–543 (2011).

McCoy AJ, Grosse-Kunstleve R., Adams PD, Win, MD, Storoni LC & Read RJ. Phaser crystallographic software. J. Appl. Cryst. 40, 658–674 (2007).

Structural studies

Demydchuk M, Hill CH, Zhou A, Bunkóczi G, Stein PE, Marchesan D, Deane JE & Read RJ. Insights into Hunter syndrome from the structure of iduronate-2-sulfatase. Nature Comm. 8, 15786 (2017).

Zhou A, Carrell RW, Murphy MP, Wei Z, Yan Y, Stanley PLD, Stein PE, Broughton Pipkin F & Read RJ. A redox switch in angiotensinogen modulates angiotensin release. Nature 468, 108–111 (2010).

Other Professional Activities

Section editor of Acta Crystallographica Section D (Structural Biology)

Member of CCP4 Executive Committee

Wellcome Trust Principal Research Fellow

Contact Details

Takes PhD students
Not available for consultancy

Affiliations

Classifications: 
Departments and institutes: