PDBsum entry 2nu8

Ligase

PDB id: 2nu8

Contents

Protein chains

287 a.a. *

388 a.a. *

Ligands

PO4 ×2

SO4 ×4

COA ×4

GOL

Waters ×510

* Residue conservation analysis

PDB id:

2nu8

Name:

Ligase

Title:

C123at mutant of e. Coli succinyl-coa synthetase

Structure:

Succinyl-coa ligase [adp-forming] subunit alpha. Chain: a, d. Synonym: succinyl-coa synthetase subunit alpha, scs-alpha. Engineered: yes. Mutation: yes. Succinyl-coa synthetase beta chain. Chain: b, e. Synonym: scs-beta. Engineered: yes

Source:

Escherichia coli. Organism_taxid: 562. Gene: sucd. Expressed in: escherichia coli. Expression_system_taxid: 562. Gene: succ.

Resolution:

2.15Å R-factor: 0.210 R-free: 0.255

Authors:

M.E.Fraser

Key ref:

E.Hidber et al. (2007). Participation of Cys123alpha of Escherichia coli succinyl-CoA synthetase in catalysis. Acta Crystallogr D Biol Crystallogr, 63, 876-884. PubMed id: 17642514 DOI: 10.1107/S0907444907029319

Date:

08-Nov-06 Release date: 24-Jul-07

Protein chains

P0AGE9  (SUCD_ECOLI) -  Succinate--CoA ligase [ADP-forming] subunit alpha from Escherichia coli (strain K12)

Seq: 289 a.a.

Struc: 287 a.a.*

Protein chains

P0A836  (SUCC_ECOLI) -  Succinate--CoA ligase [ADP-forming] subunit beta from Escherichia coli (strain K12)

Seq: 388 a.a.

Struc: 388 a.a.

* PDB and UniProt seqs differ at 1 residue position (black cross)

Enzyme reactions

• Enzyme class: Chains A, B, D, E: E.C.6.2.1.5 - succinate--CoA ligase (ADP-forming).
• Reaction: succinate + ATP + CoA = succinyl-CoA + ADP + phosphate

succinate + ATP (Bound ligand (Het Group name = COA) corresponds exactly) + CoA (Bound ligand (Het Group name = GOL) matches with 40.00% similarity) = succinyl-CoA + ADP (Bound ligand (Het Group name = PO4) corresponds exactly) + phosphate

Molecule diagrams generated from .mol files obtained from the KEGG ftp site

reference

DOI no: 10.1107/S0907444907029319

Acta Crystallogr D Biol Crystallogr 63:876-884 (2007)

PubMed id: 17642514

Participation of Cys123alpha of Escherichia coli succinyl-CoA synthetase in catalysis.

E.Hidber, E.R.Brownie, K.Hayakawa, M.E.Fraser.

ABSTRACT

Succinyl-CoA synthetase has a highly conserved cysteine residue, Cys123alpha in the Escherichia coli enzyme, that is located near the CoA-binding site and the active-site histidine residue. To test whether the succinyl moiety of succinyl-CoA is transferred to the thiol of Cys123alpha as part of the catalytic mechanism, this residue was mutated to alanine, serine, threonine and valine. Each mutant protein was catalytically active, although less active than the wild type. This proved that the specific formation of a thioester bond with Cys123alpha is not part of the catalytic mechanism. To understand why the mutations affected catalysis, the crystal structures of the four mutant proteins were determined. The alanine mutant showed no structural changes yet had reduced activity, suggesting that the size of the cysteine is important for optimal activity. These results explain why this cysteine residue is conserved in the sequences of succinyl-CoA synthetases from different sources.

Selected figure(s)

Figure 4.
Figure 4 Stereoview showing part of the C123 V mutant, including the phosphohistidine loop. For comparison, the model of the C123 T mutant protein from the tetragonal crystal form is superposed. Shading is according to atom type, as in Fig. 1-, but the C atoms for the C123 T mutant are cyan. Dotted lines show hydrogen-bonding interactions present in the C123 T mutant but not in the C123 V mutant.

Figure 5.
Figure 5 Stereoviews of the structures near residue 123 of the -subunit showing the effects of disorder of the phosphohistidine loop. The structures are shown as ball-and-stick drawings shaded according to atom type as in Fig. 1-. Dashed lines show hydrogen-bonding interactions. (a) -Subunit of T. thermophilus SCS (PDB code 1oi7 ). (b) Wild-type E. coli SCS (Fraser et al., 2002[Fraser, M. E., Joyce, M. A., Ryan, D. G. & Wolodko, W. T. (2002). Biochemistry, 41, 537-546.]). (c) C123 V mutant protein.

The above figures are reprinted by permission from the IUCr: Acta Crystallogr D Biol Crystallogr (2007, 63, 876-884) copyright 2007.

Figures were selected by the author.