Native SAD is maturing.

John P Rose,Bi-Cheng Wang,Manfred S Weiss

doi:10.1107/s2052252515008337

Abstract

Native SAD phasing uses the anomalous scattering signal of light atoms in the crystalline, native samples of macromolecules collected from single-wavelength X-ray diffraction experiments. These atoms include sodium, magnesium, phosphorus, sulfur, chlorine, potassium and calcium. Native SAD phasing is challenging and is critically dependent on the collection of accurate data. Over the past five years, advances in diffraction hardware, crystallographic software, data-collection methods and strategies, and the use of data statistics have been witnessed which allow 'highly accurate data' to be routinely collected. Today, native SAD sits on the verge of becoming a 'first-choice' method for both de novo and molecular-replacement structure determination. This article will focus on advances that have caught the attention of the community over the past five years. It will also highlight both de novo native SAD structures and recent structures that were key to methods development.

Highlights

Native SAD phasing uses the anomalous scattering signal of atoms in the crystalline, native samples of macromolecules collected from single-wavelength X-ray diffraction experiments
With the exception of metalloproteins, native SAD phasing is critically dependent on accurately recording the weak anomalous scattering signal from light atoms such as sulfur, phosphorus, chlorine, potassium, calcium and magnesium present in the crystallized sample
Over the past decade similar workshops and schools have been given at various locations worldwide, including MS 40: sulfur SAD (S-SAD) and Other Applications of Soft X-rays in MX at the 2014 Congress of the International Union of Crystallography in Montreal and the fifth Winter School on Soft X-rays in Macromolecular Crystallography held at the University of Georgia in March 2015

Summary

Introduction

Native SAD phasing uses the anomalous scattering signal of atoms in the crystalline, native samples of macromolecules collected from single-wavelength X-ray diffraction experiments. With the exception of metalloproteins, native SAD phasing is critically dependent on accurately recording the weak anomalous scattering signal from light atoms such as sulfur, phosphorus, chlorine, potassium, calcium and magnesium present in the crystallized sample. That the Bence–Jones protein Rhe (two S atoms in 113 resi- (52) of the native SAD structures reported (i.e. structures that dues) could be successfully phased using only the anomalous have a Protein Data Bank ID) have been determined from scattering signal from a single disulfide bond. The removal of these bottlenecks was key to the structure determination of the second de novo S-SAD structure 20 years later. The first data sets from the beamline are expected in early 2015

Detectors

Sample preparation

Data collection

Data processing

Phasing and structure solution

Building community

Findings

Discussion