PMID- 9698564
OWN - NLM
STAT- MEDLINE
DCOM- 19980916
LR  - 20071114
IS  - 0022-2836 (Print)
IS  - 0022-2836 (Linking)
VI  - 281
IP  - 3
DP  - 1998 Aug 21
TI  - A pH-dependent stabilization of an active site loop observed from low and high pH 
      crystal structures of mutant monomeric glycinamide ribonucleotide transformylase 
      at 1.8 to 1.9 A.
PG  - 485-99
AB  - A mutation in the dimer interface of Escherichia coli glycinamide ribonucleotide 
      transformylase (GarTfase) disrupts the observed pH-dependent association of the 
      wild-type enzyme, but has no observable effect on the enzyme activity. Here, we 
      assess whether a pH effect on the enzyme's conformation is sufficient by itself 
      to explain the pH-dependence of the GarTfase reaction. A pH-dependent 
      conformational change is observed between two high-resolution crystal structures 
      of the Glu70Ala mutant GarTfase at pH 3.5 (1.8 A) and 7.5 (1.9 A). Residues 110 
      to 131 in GarTfase undergo a transformation from a disordered loop at pH 3.5, 
      where the enzyme is inactive, to an ordered loop-helix structure at pH 7.5, where 
      the enzyme is active. The ordering of this flexible loop-helix has a direct 
      effect on catalytic residues in the active site, binding of the folate cofactor 
      and shielding of the active site from solvent. A main-chain carbonyl oxygen atom 
      from Tyr115 in the ordered loop forms a hydrogen bond with His108, and thereby 
      provides electronic and structural stabilization of this key active site residue. 
      Kinetic data indicate that the pKa of His108 is in fact raised to 9. 2. The loop 
      movement can be correlated with elevation of the His pKa, but with further 
      stabilization, probably from Asp144, after the binding of folate cofactor. 
      Leu118, also in the loop, becomes positioned near the p-amino benzoic acid 
      binding site, providing additional hydrophobic interactions with the cofactor 
      10-formyl tetrahydrofolate. Thus, the pH-dependence of the enzyme activity 
      appears to arise from local active site rearrangements and not from differences 
      due to monomer-dimer association.
CI  - Copyright 1998 Academic Press.
FAU - Su, Y
AU  - Su Y
AD  - Department of Molecular Biology and The Skaggs Institute for Chemical Biology, 
      The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA, 
      92093-0359, USA.
FAU - Yamashita, M M
AU  - Yamashita MM
FAU - Greasley, S E
AU  - Greasley SE
FAU - Mullen, C A
AU  - Mullen CA
FAU - Shim, J H
AU  - Shim JH
FAU - Jennings, P A
AU  - Jennings PA
FAU - Benkovic, S J
AU  - Benkovic SJ
FAU - Wilson, I A
AU  - Wilson IA
LA  - eng
SI  - PDB/2GAR
SI  - PDB/3GAR
GR  - GM54038/GM/NIGMS NIH HHS/United States
GR  - P01-CA63536/CA/NCI NIH HHS/United States
GR  - T32 MH19185/MH/NIMH NIH HHS/United States
GR  - etc.
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PT  - Research Support, U.S. Gov't, P.H.S.
PL  - Netherlands
TA  - J Mol Biol
JT  - Journal of molecular biology
JID - 2985088R
RN  - 0 (Coenzymes)
RN  - EC 2.1.2.- (Hydroxymethyl and Formyl Transferases)
RN  - EC 2.1.2.2 (Phosphoribosylglycinamide Formyltransferase)
SB  - IM
MH  - Binding Sites
MH  - Coenzymes
MH  - Crystallography, X-Ray/methods
MH  - Escherichia coli/*enzymology
MH  - Hydrogen-Ion Concentration
MH  - Hydroxymethyl and Formyl Transferases/*chemistry/genetics
MH  - Kinetics
MH  - Models, Molecular
MH  - Phosphoribosylglycinamide Formyltransferase
MH  - Point Mutation
MH  - *Protein Conformation
EDAT- 1998/08/12 00:00
MHDA- 1998/08/12 00:01
CRDT- 1998/08/12 00:00
PHST- 1998/08/12 00:00 [pubmed]
PHST- 1998/08/12 00:01 [medline]
PHST- 1998/08/12 00:00 [entrez]
AID - S0022-2836(98)91931-3 [pii]
AID - 10.1006/jmbi.1998.1931 [doi]
PST - ppublish
SO  - J Mol Biol. 1998 Aug 21;281(3):485-99. doi: 10.1006/jmbi.1998.1931.