PDBsum entry 1zdf

Transferase

PDB id: 1zdf

Contents

Protein chain

258 a.a. *

Ligands

SO4

UPG

Metals

_MN

Waters ×101

* Residue conservation analysis

PDB id:

1zdf

Name:

Transferase

Title:

Ser162 mutant of glycogenin complexed with udp-glucose and manganese

Structure:

Glycogenin-1. Chain: a. Engineered: yes. Mutation: yes

Source:

Oryctolagus cuniculus. Rabbit. Organism_taxid: 9986. Gene: gyg, gyg1. Expressed in: escherichia coli bl21. Expression_system_taxid: 511693.

Biol. unit:

Dimer (from PDB file)

Resolution:

2.45Å R-factor: 0.199 R-free: 0.217

Authors:

T.D.Hurley,S.L.Stout,E.Miner,J.Zhou,P.J.Roach

Key ref:

T.D.Hurley et al. (2005). Requirements for catalysis in mammalian glycogenin. J Biol Chem, 280, 23892-23899. PubMed id: 15849187 DOI: 10.1074/jbc.M502344200

Date:

14-Apr-05 Release date: 26-Apr-05

Protein chain

P13280  (GLYG_RABIT) -  Glycogenin-1 from Oryctolagus cuniculus

Seq: 333 a.a.

Struc: 258 a.a.*

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

Enzyme reactions

• Enzyme class: E.C.2.4.1.186 - glycogenin glucosyltransferase.
• Reaction:
  1. L-tyrosyl-[glycogenin] + UDP-alpha-D-glucose = alpha-D-glucosyl-L- tyrosyl-[glycogenin] + UDP + H⁺
  2. [1,4-alpha-D-glucosyl](n)-L-tyrosyl-[glycogenin] + UDP-alpha-D- glucose = [1,4-alpha-D-glucosyl](n+1)-L-tyrosyl-[glycogenin] + UDP + H⁺

L-tyrosyl-[glycogenin] + UDP-alpha-D-glucose (Bound ligand (Het Group name = UPG) corresponds exactly) = alpha-D-glucosyl-L- tyrosyl-[glycogenin] + UDP + H(+)

[1,4-alpha-D-glucosyl](n)-L-tyrosyl-[glycogenin] + UDP-alpha-D- glucose (Bound ligand (Het Group name = UPG) corresponds exactly) = [1,4-alpha-D-glucosyl](n+1)-L-tyrosyl-[glycogenin] + UDP + H(+)

• Cofactor: Mn(2+)

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

reference

DOI no: 10.1074/jbc.M502344200

J Biol Chem 280:23892-23899 (2005)

PubMed id: 15849187

Requirements for catalysis in mammalian glycogenin.

T.D.Hurley, S.Stout, E.Miner, J.Zhou, P.J.Roach.

ABSTRACT

Glycogenin is a glycosyltransferase that functions as the autocatalytic initiator for the synthesis of glycogen in eukaryotic organisms. Prior structural work identified the determinants responsible for the recognition and binding of UDP-glucose and the catalytic manganese ion and implicated two aspartic acid residues in the reaction mechanism for self-glucosylation. We examined the effects of substituting asparagine and serine for the aspartic acid residues at positions 159 and 162. We also examined whether the truncation of the protein at residue 270 (delta270) was compatible with its structural integrity and its functional role as the initiator for glycogen synthesis. The truncated form of the enzyme was indistinguishable from the wild-type enzyme by all measures of activity and could support glycogen accumulation in a glycogenin-deficient yeast strain. Substitution of aspartate 159 by either serine or asparagine eliminated self-glucosylation and reduced trans-glucosylation activity by at least 260-fold but only reduced UDP-glucose hydrolytic activity by 4-14-fold. Substitution of aspartate 162 by either serine or asparagine eliminated self-glucosylation activity and reduced UDP-glucose hydrolytic activity by at least 190-fold. The trans-glucosylation of maltose was reduced to undetectable levels in the asparagine 162 mutant, whereas the serine 162 enzyme showed only an 18-30-fold reduction in its ability to trans-glucosylate maltose. These data support a role for aspartate 162 in the chemical step for the glucosyltransferase reaction and a role for aspartate 159 in binding and activating the acceptor molecule.

Selected figure(s)

Figure 1.
FIG. 1. JZ4-a cells stained for glycogen accumulation using iodine vapor. A, JZ4-a cells transformed only with wild-type glycogenin-1. B, JZ4-a cells transformed only with 194F glycogenin-1. C, JZ4-a cells transformed with both 194F-glycogenin-1 and 271-103A glycogenin-1.

Figure 2.
FIG. 2. Subunit relationships among glycogenin crystal forms. A, tetrameric association formed by crystallographic contacts between the dimers that comprise the asymmetric unit in the 270 crystals. Each subunit is colored differently, and the position of Tyr-194 in each subunit is highlighted using purple space-filling atoms, and the position of the bound UDP molecules is highlighted using blue space-filling atoms. B, alignment of the dimer formed by one of the crystallographic axes in the I222 space group (red) with the dimer of the 270 enzyme that comprises the asymmetric unit of the P6[4] space group (yellow). For this figure only the respective "A" subunits were aligned. The red arrow indicates the amount of additional rotation required to align the "B" subunits using the molecules oriented in this manner. Figure was prepared using the programs MOLSCRIPT (38) and Raster3D (39, 40).

The above figures are reprinted from an Open Access publication published by the ASBMB: J Biol Chem (2005, 280, 23892-23899) copyright 2005.

Figures were selected by an automated process.