PDBsum entry 4eab

Transferase

PDB id: 4eab

Contents

Protein chain

210 a.a.

Ligands

JB2

COA

Metals

_CL

_NA

Waters ×229

PDB id:

4eab

Name:

Transferase

Title:

X-ray crystal structure of the h141a mutant of gdp-perosamine n-acetyl transferase from caulobacter crescentus in complex with coa and gdp- perosamine

Structure:

Perosamine n-acetyltransferase. Chain: a. Synonym: perb, hexapeptide transferase family protein. Engineered: yes. Mutation: yes

Source:

Caulobacter vibrioides. Organism_taxid: 155892. Gene: cc_1011, wbqr. Expressed in: escherichia coli. Expression_system_taxid: 469008.

Resolution:

1.35Å R-factor: 0.188 R-free: 0.204

Authors:

J.B.Thoden,L.A.Reinhardt,P.D.Cook,P.Menden,W.W.Cleland,H.M.Holden

Key ref:

J.B.Thoden et al. (2012). Catalytic mechanism of perosamine N-acetyltransferase revealed by high-resolution X-ray crystallographic studies and kinetic analyses. Biochemistry, 51, 3433-3444. PubMed id: 22443398

Date:

22-Mar-12 Release date: 04-Apr-12

Protein chain

O85353  (O85353_CAUVI) -  Putative acetyltransferase from Caulobacter vibrioides

Seq: 215 a.a.

Struc: 210 a.a.*

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

Enzyme reactions

• Enzyme class: E.C.2.3.1.- - ?????

Biochemistry 51:3433-3444 (2012)

PubMed id: 22443398

Catalytic mechanism of perosamine N-acetyltransferase revealed by high-resolution X-ray crystallographic studies and kinetic analyses.

J.B.Thoden, L.A.Reinhardt, P.D.Cook, P.Menden, W.W.Cleland, H.M.Holden.

ABSTRACT

N-Acetylperosamine is an unusual dideoxysugar found in the O-antigens of some Gram-negative bacteria, including the pathogenic Escherichia coli strain O157:H7. The last step in its biosynthesis is catalyzed by PerB, an N-acetyltransferase belonging to the left-handed β-helix superfamily of proteins. Here we describe a combined structural and functional investigation of PerB from Caulobacter crescentus. For this study, three structures were determined to 1.0 Å resolution or better: the enzyme in complex with CoA and GDP-perosamine, the protein with bound CoA and GDP-N-acetylperosamine, and the enzyme containing a tetrahedral transition state mimic bound in the active site. Each subunit of the trimeric enzyme folds into two distinct regions. The N-terminal domain is globular and dominated by a six-stranded mainly parallel β-sheet. It provides most of the interactions between the protein and GDP-perosamine. The C-terminal domain consists of a left-handed β-helix, which has nearly seven turns. This region provides the scaffold for CoA binding. On the basis of these high-resolution structures, site-directed mutant proteins were constructed to test the roles of His 141 and Asp 142 in the catalytic mechanism. Kinetic data and pH-rate profiles are indicative of His 141 serving as a general base. In addition, the backbone amide group of Gly 159 provides an oxyanion hole for stabilization of the tetrahedral transition state. The pH-rate profiles are also consistent with the GDP-linked amino sugar substrate entering the active site in its unprotonated form. Finally, for this investigation, we show that PerB can accept GDP-3-deoxyperosamine as an alternative substrate, thus representing the production of a novel trideoxysugar.