Increasing the X-ray Diffraction Power of Protein Crystals by Dehydration: The Case of Bovine Serum Albumin and a Survey of Literature Data

Abstract

Serum albumin is one of the most widely studied proteins. It is the most abundant protein in plasma with a typical concentration of 5 g/100 mL and the principal transporter of fatty acids in plasma. While the crystal structures of human serum albumin (HSA) free and in complex with fatty acids, hemin, and local anesthetics have been characterized, no crystallographic models are available on bovine serum albumin (BSA), presumably because of the poor diffraction power of existing hexagonal BSA crystals. Here, the crystallization and diffraction data of a new BSA crystal form, obtained by the hanging drop method using MPEG 5K as precipitating agent, are presented. The crystals belong to space group C2, with unit-cell parameters a = 216.45 Å, b = 44.72 Å, c = 140.18 Å, β = 114.5°. Dehydration was found to increase the diffraction limit of BSA crystals from ~8 Å to 3.2 Å, probably by improving the packing of protein molecules in the crystal lattice. These results, together with a survey of more than 60 successful cases of protein crystal dehydration, confirm that it can be a useful procedure to be used in initial screening as a method of improving the diffraction limits of existing crystals.

Dramatic Improvement of Crystals of Large RNAs by Cation Replacement and Dehydration

“…Several studies have reported on the utility of controlled dehydration for improvement of the diffraction properties of protein crystals, and investigated how protein molecules reorganized to form improved packing contacts (Deng et al, 2012; Heras and Martin, 2005; Russo Krauss et al, 2012). Using a similar approach, we have previously found that brief soaking in a high osmolarity solution of crystals of the glmS ribozyme-riboswitch improved their diffraction limit (Klein and Ferré-D’Amaré, 2009; Klein et al, 2007) from 3.0 Å to 1.7 Å.…”

An Overview of Biological Macromolecule Crystallization

“…Post-crystallization methods are more crucial for large proteins and macromolecular complexes since a new, and possibly better diffracting, crystal form is usually more difficult to obtain compared to small protein targets. The most used post-crystallization methods for improvement of the diffraction properties include soaking, cross-linking, annealing [261,262] and controlled dehydration [263,264]. Soaking of protein crystals with their product analogs, inhibitors, or strong ligands can be used to stabilize protein conformation and increase the crystalline lattice regularity.…”

“…In the fortunate cases, this procedure induces an internal rearrangement of the crystal that results in a significantly improved quality of the diffraction data. The various techniques for dehydrating crystals and the proteins whose diffraction pattern was improved through this procedure have been summarized in a recent paper [263]. To assure the control and reproducibility of dehydration, a number of dedicated devices have been produced [267–270], some of which monitor in real time the X-ray diffraction pattern of the crystal [271–273].…”

Protein Recognition of Gold-Based Drugs: 3D Structure of the Complex Formed When Lysozyme Reacts with Aubipyc

“…This methodology can yield improved low-temperature crystalline order, 17 higher resolution of X-ray diffraction, 18 and reduced mosaicity.…”