The role of Co²⁺in the crystallization of human SENP1 and comments on the limitations of automated refinement protocols

Abstract

PDB References: SENP1, 2xph; 2xre.Metal ions often stabilize intermolecular contacts between macromolecules, thereby promoting crystallization. When interpreting a medium-resolution electron-density map of the catalytic domain of human sentrin-specific protease 1 (SENP1), a strong feature indicative of an ordered divalent cation was noted. This was assigned as Co 2+ , an essential component of the crystallization mixture. The ion displays tetrahedral coordination by Glu430 and His640 from one molecule and the corresponding residues from a symmetry-related molecule. Analysis of the data derived from a previous structure of SENP1 suggested that Co 2+ had been overlooked and re-refinement supported this conclusion. Highthroughput automated re-refinement protocols also failed to mark the Co 2+ position, supporting the requirement for the incorporation of as much information as possible to enhance the value of such protocols.

Automated identification of elemental ions in macromolecular crystal structures

“…Similar errors have been observed in published structures (e.g. Rimsa et al, 2011). Especially at less than atomic resolution, refinement as water will push apart an ion and its ligands somewhat, making the valence-bond analysis less reliable.…”

“…If crystals are sufficiently plentiful and/or radiation-tolerant, it may also be worthwhile collecting an additional lower-resolution data set at a longer wavelength. Furthermore, we strongly encourage the deposition of the separate Friedel pairs (or, better yet, the unmerged intensities) in the Protein Data Bank upon publication, as these provide essential information about the experiment and may lead to future improvements in both the development of crystallographic software and in the deposited structures themselves (Joosten et al, 2009(Joosten et al, , 2012Rimsa et al, 2011;Afonine et al, 2012).…”

Structures ofPseudomonas aeruginosaβ-ketoacyl-(acyl-carrier-protein) synthase II (FabF) and a C164Q mutant provide templates for antibacterial drug discovery and identify a buried potassium ion and a ligand-binding site that is an artefact of the crystal form

“…A striking and unusual observation is the identification of a buried cation approximately 8 Å from the catalytic Cys164 in all three structures. Care was taken in the assignment of this ion based on correlation of difference density (F o À F c ) levels with the number of electrons (Fyfe et al, 2009;Rimsa et al, 2011). The exclusion of all eight S atoms in PaFabF produced positive peaks in the difference density map with an average height of 18.6.…”