Lysine methylation as a routine rescue strategy for protein crystallization

Abstract

Crystallization remains a critical step in X-ray structure determination. Because it is not generally possible to rationally predict crystallization conditions, commercial screens have been developed which sample a wide range of crystallization space. While this approach has proved successful in many cases, a significant number of proteins fail to crystallize despite being soluble and monodispersed. It is established that chemical modification can facilitate the crystallization of otherwise intractable proteins. Here we describe a method for the reductive methylation of lysine residues which is simple, inexpensive, and efficient, and report on its application to ten proteins. We describe the effect of methylation on the physico-chemical properties of these proteins, and show that it led to diffraction-quality crystals from four proteins and structures for three that had hitherto proved refractory to crystallization. The method is suited to both low- and high-throughput laboratories.

Figure 1

Examples of Methylated Lysines from MVE Methyltransferase (A) An example of the interaction between a methylated lysine and a glutamate residue at a crystal contact. An overview of the packing of molecules of MVE methyltransferase in one of the three crystal forms solved (space group P2₁, a = 46.4 Å, b = 68.3 Å, c = 82.0 Å, β = 98.4°) is drawn in the top panel, with the two molecules in the crystallographic asymmetric unit colored red and green, respectively. The close-up in the bottom panel shows the hydrophobic packing of the aliphatic components of the side chains lysine 177 and glutamate 166. (B) An example of well-defined electron density for a methylated lysine (K15) involved in a crystal contact with the main chain of a neighboring molecule. The figure was produced using PyMOL (http://www.pymol.org).