PDBsum entry 1vjl

Unknown function

PDB id

1vjl

Contents

			Protein chains

					143 a.a. *

			Ligands

					UNL ×2

			Metals

					_CL ×2

			Waters ×164

* Residue conservation analysis

References listed in PDB file

Key reference

Title

On the use of dxms to produce more crystallizable proteins: structures of the t. Maritima proteins tm0160 and tm1171.

Authors

G.Spraggon, D.Pantazatos, H.E.Klock, I.A.Wilson, V.L.Woods, S.A.Lesley.

Ref.

Protein Sci, 2004, 13, 3187-3199. [DOI no: 10.1110/ps.04939904]

PubMed id

15557262

Abstract

The structure of two Thermotoga maritima proteins, a conserved hypothetical protein (TM0160) and a transcriptional regulator (TM1171), have now been determined at 1.9 A and 2.3 A resolution, respectively, as part of a large-scale structural genomics project. Our first efforts to crystallize full-length versions of these targets were unsuccessful. However, analysis of the recombinant purified proteins using the technique of enhanced amide hydrogen/deuterium exchange mass spectroscopy (DXMS) revealed substantial regions of rapid amide deuterium hydrogen exchange, consistent with flexible regions of the structures. Based on these exchange data, truncations were designed to selectively remove the disordered C-terminal regions, and the resulting daughter proteins showed greatly enhanced crystallizability. Comparative DXMS analysis of full-length protein versus truncated forms demonstrated complete and exact preservation of the exchange rate profiles in the retained sequence, indicative of conservation of the native folded structure. This study presents the first structures produced with the aid of the DXMS method for salvaging intractable crystallization targets. The structure of TM0160 represents a new fold and highlights the use of this approach where any prior structural knowledge is absent. The structure of TM1171 represents an example where the lack of a substrate/cofactor may impair crystallization. The details of both structures are presented and discussed.



	Figure 2. Figure 2. Structure of TM0160. (A) Topology diagram of the overall fold of TM0160. The long mixed -sheet is shaded cyan. The dimerization helix (H2) is shaded yellow, while the highly mobile C-terminal epitope tag helix (H5) in molecule A is shaded red; all other helices are shaded green. The picture was generated by TOPS (Westhead et al. 1999) and Topdraw (Bond 2003). (B) Stereo diagram of the TM0160 monomer generated by VMD (Humphrey et al. 1996). C atom numbering is every 20 residues. (C) Two orthogonal ribbon diagram representation of the TM0160 dimer. The interchain disulfide is depicted in a ball-and-stick representation and sits on the molecular twofold displayed as an arrow in the top diagram and as an oval in the bottom. The ribbon is colored from blue to green in subunit A and green to red in subunit B. The figure was generated using Bobscript (Kraulis 1991; Esnouf 1997) and Raster3d (Meritt and Murphy 1994). (D) Representative 2Fo-Fc electron density. The electron density of the region around the molecular twofold axis details the interchain disulfide bond. The electron density map is contoured at 1.5 standard deviations above the mean.		Figure 5. Figure 5. Comparison of TM1171 with E. coli transcription regulator. (A) Ribbon diagram of E. coli transcription regulator in complex with its DNA substrate (Parkinson et al. 1996). CRP domain bound to DNA molecule A of the dimer is colored cyan and the other is colored yellow. Regions defined by SEG to be disordered are shaded red, while those for DXMS are shaded green (Fig. 1 Go-). DNA is represented by ball-and-stick. The figure was generated with Bobscript (Kraulis 1991; Esnouf 1997) and Raster3d (Meritt and Murphy 1994). (B) Superposition of TM1171 cNTP domain with its counterpart in E. coli (PDB code 1RUN [PDB] ). TM1171 is colored red and 1RUN [PDB] is colored yellow. The overall rmsd between the two domains is 1.74 � over 111 aligned C residues; the dimerization helix is rotated relative to its counterpart in 1RUN [PDB] by about 20�.

PROCHECK

	Headers