PMID- 10587441
OWN - NLM
STAT- MEDLINE
DCOM- 20000110
LR  - 20190613
IS  - 0006-2960 (Print)
IS  - 0006-2960 (Linking)
VI  - 38
IP  - 49
DP  - 1999 Dec 7
TI  - An asparagine-phenylalanine substitution accounts for catalytic differences
      between hGSTM3-3 and other human class mu glutathione S-transferases.
PG  - 16187-94
AB  - The hGSTM3 subunit, which is preferentially expressed in germ-line cells, has the
      greatest sequence divergence among the human mu class glutathione S-transferases.
      To determine a structural basis for the catalytic differences between hGSTM3-3
      and other mu class enzymes, chimeric proteins were designed by modular
      interchange of the divergent C-terminal domains of hGSTM3 and hGSTM5 subunits.
      Replacement of 24 residues of the C-terminal segment of either subunit produced
      chimeric enzymes with catalytic properties that reflected those of the wild-type 
      enzyme from which the C-terminus had been derived. Deletion of the tripeptide
      C-terminal extension found only in the hGSTM3 subunit had no effect on catalysis.
      The crystal structure determined for a ligand-free hGSTM3 subunit indicates that 
      an Asn212 residue of the C-terminal domain is near a hydrophobic cluster of side 
      chains formed in part by Ile13, Leu16, Leu114, Ile115, Tyr119, Ile211, and
      Trp218. Accordingly, a series of point mutations were introduced into the hGSTM3 
      subunit, and it was indeed determined that a Y119F mutation considerably enhanced
      the turnover rate of the enzyme for nucleophilic aromatic substitution reactions.
      A more striking effect was observed for a double mutant (Y119F/N212F) which had a
      k(cat)/K(m)(CDNB) value of 7.6 x 10(5) s(-)(1) M(-)(1) as compared to 4.9 x 10(3)
      s(-)(1) M(-)(1) for the wild-type hGSTM3-3 enzyme. The presence of a polar Asn212
      in place of a Phe residue found in the cognate position of other mu class
      glutathione S-transferases, therefore, has a marked influence on catalysis by
      hGSTM3-3.
FAU - Patskovsky, Y V
AU  - Patskovsky YV
AD  - Department of Biochemistry, Albert Einstein College of Medicine, Bronx, New York 
      10461, USA.
FAU - Patskovska, L N
AU  - Patskovska LN
FAU - Listowsky, I
AU  - Listowsky I
LA  - eng
SI  - PDB/3GTU
GR  - CA 42448/CA/NCI NIH HHS/United States
PT  - Comparative Study
PT  - Journal Article
PT  - Research Support, U.S. Gov't, P.H.S.
PL  - United States
TA  - Biochemistry
JT  - Biochemistry
JID - 0370623
RN  - 0 (Recombinant Fusion Proteins)
RN  - 2497-91-8 (2-carboxy-4,6-dinitrochlorobenzene)
RN  - 47E5O17Y3R (Phenylalanine)
RN  - 7006-34-0 (Asparagine)
RN  - EC 2.5.1.18 (Glutathione Transferase)
RN  - GE3IBT7BMN (Dinitrochlorobenzene)
SB  - IM
MH  - Amino Acid Sequence
MH  - Amino Acid Substitution/*genetics
MH  - Asparagine/*genetics/metabolism
MH  - Binding Sites/genetics
MH  - Catalysis
MH  - Crystallography, X-Ray
MH  - Dinitrochlorobenzene/analogs & derivatives/metabolism
MH  - Glutathione Transferase/chemistry/classification/*genetics/*metabolism
MH  - Humans
MH  - Hydrogen-Ion Concentration
MH  - Kinetics
MH  - Molecular Sequence Data
MH  - Mutagenesis, Site-Directed
MH  - Phenylalanine/*genetics/metabolism
MH  - Point Mutation
MH  - Recombinant Fusion Proteins/chemistry/genetics/metabolism
EDAT- 1999/12/10 00:00
MHDA- 1999/12/10 00:01
CRDT- 1999/12/10 00:00
PHST- 1999/12/10 00:00 [pubmed]
PHST- 1999/12/10 00:01 [medline]
PHST- 1999/12/10 00:00 [entrez]
AID - bi991714t [pii]
AID - 10.1021/bi991714t [doi]
PST - ppublish
SO  - Biochemistry. 1999 Dec 7;38(49):16187-94. doi: 10.1021/bi991714t.