DB code: D00273

RLCP classification 8.131.581400.399 : Isomerization
8.113.580000.386 : Isomerization
CATH domain 3.30.390.10 : Enolase-like; domain 1
3.20.20.120 : TIM Barrel Catalytic domain
E.C. 5.1.2.2
CSA 1mdr
M-CSA 1mdr
MACiE M0187

CATH domain Related DB codes (homologues)
3.20.20.120 : TIM Barrel D00261 D00282 D00283
3.30.390.10 : Enolase-like; domain 1 D00261 D00282 D00283

Uniprot Enzyme Name
UniprotKB Protein name Synonyms Pfam
P11444 Mandelate racemase
MR
EC 5.1.2.2
PF01188 (MR_MLE)
PF02746 (MR_MLE_N)
[Graphical View]

KEGG enzyme name
mandelate racemase

UniprotKB: Accession Number Entry name Activity Subunit Subcellular location Cofactor
P11444 MANR_PSEPU (S)-mandelate = (R)-mandelate. Homooctamer. Divalent metal ions. Magnesium seems to be the preferred ion.

KEGG Pathways
Map code Pathways E.C.
MAP00362 Benzoate degradation via hydroxylation
MAP00622 Toluene and xylene degradation

Compound table
Cofactors Substrates Products Intermediates
KEGG-id C00305 C01984 C01983 I00074
E.C.
Compound Magnesium (S)-Mandelate (R)-Mandelate Phenylethene-1,2-triol
Type divalent metal (Ca2+, Mg2+) aromatic ring (only carbon atom),carbohydrate,carboxyl group aromatic ring (only carbon atom),carbohydrate,carboxyl group
ChEBI 18420
32800
17656
PubChem 888
439616
11914
1dtnA01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Unbound Unbound Unbound Unbound
1mdlA01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Unbound Unbound Unbound Unbound
1mdrA01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Unbound Unbound Unbound Unbound
1mnsA01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Unbound Unbound Unbound Unbound
1mraA01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Unbound Unbound Unbound Unbound
2mnrA01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Unbound Unbound Unbound Unbound
1dtnA02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Bound:_MG Analogue:APG Unbound Unbound
1mdlA02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Bound:_MG Bound:SMN Unbound Unbound
1mdrA02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Bound:_MG Analogue:APG Unbound Unbound
1mnsA02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Bound:_MG Analogue:APG Unbound Unbound
1mraA02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Bound:_MG Analogue:APG Unbound Unbound
2mnrA02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain Analogue:_MN Unbound Unbound Unbound

Reference for Active-site residues
resource references E.C.
Swiss-prot;P11444 & literature [4], [9], [10], [11], [12], [14]

Active-site residues
PDB Catalytic residues Cofactor-binding residues Modified residues Main-chain involved in catalysis Comment
1dtnA01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain
1mdlA01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain
1mdrA01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain
1mnsA01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain
1mraA01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain
2mnrA01 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain
1dtnA02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain LYS 164;LYS 166;ASP 270;HIS 297; ASP 195;GLU 221;GLU 247(Magnesium binding) mutant E317Q
1mdlA02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain LYS 164;;ASP 270;HIS 297;GLU 317 ASP 195;GLU 221;GLU 247(Magnesium binding) mutant K166R
1mdrA02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain LYS 164;LYS 166;ASP 270;HIS 297;GLU 317 ASP 195;GLU 221;GLU 247(Magnesium binding)
1mnsA02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain LYS 164;LYS 166;ASP 270;HIS 297;GLU 317 ASP 195;GLU 221;GLU 247(Magnesium binding) LYS 166(APG binding)
1mraA02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain LYS 164;LYS 166;;HIS 297;GLU 317 ASP 195;GLU 221;GLU 247(Magnesium binding) mutant D270N
2mnrA02 Pdbj logo s Rasmollogo id Rasmollogo chain Mmcif id Mmcif chain LYS 164;LYS 166;ASP 270;HIS 297;GLU 317 ASP 195;GLU 221;GLU 247(Magnesium binding)

References for Catalytic Mechanism
References Sections No. of steps in catalysis
[9]
Scheme I
[10]
Fig.6, p.1889
[11]
p.2795-2796
[12]
Fig.2, p.2782-2783
[13]
p.16495-16496
[14]
p.5665-5667
[15]
Fig.7, p.1652-1654
[17]
p.25532
[18]
Fig.1, p.10400-10401
[19]
Scheme 1, p.13332-13334
[22]
p.152-153
[23]
Scheme 1

References
[1]
Resource
Comments
Medline ID
PubMed ID 3132459
Journal J Biol Chem
Year 1988
Volume 263
Pages 9268-70
Authors Neidhart DJ, Powers VM, Kenyon GL, Tsou AY, Ransom SC, Gerlt JA, Petsko GA
Title Preliminary x-ray data on crystals of mandelate racemase.
Related PDB
Related UniProtKB
[2]
Resource
Comments
Medline ID
PubMed ID 2099737
Journal Biochem Soc Symp
Year 1990
Volume 57
Pages 135-41
Authors Neidhart DC, Howell PL, Petsko GA, Gerlt JA, Kozarich JW, Powers VM, Kenyon GL
Title Restructuring catalysis in the mandelate pathway.
Related PDB
Related UniProtKB
[3]
Resource
Comments SIMILARITY TO MLE.
Medline ID 91015392
PubMed ID 2215699
Journal Nature
Year 1990
Volume 347
Pages 692-4
Authors Neidhart DJ, Kenyon GL, Gerlt JA, Petsko GA
Title Mandelate racemase and muconate lactonizing enzyme are mechanistically distinct and structurally homologous.
Related PDB
Related UniProtKB P11444
[4]
Resource
Comments MUTAGENESIS OF HIS-297.
Medline ID 91369941
PubMed ID 1909893
Journal Biochemistry
Year 1991
Volume 30
Pages 9274-81
Authors Landro JA, Kallarakal AT, Ransom SC, Gerlt JA, Kozarich JW, Neidhart DJ, Kenyon GL
Title Mechanism of the reaction catalyzed by mandelate racemase. 3. Asymmetry in reactions catalyzed by the H297N mutant.
Related PDB
Related UniProtKB P11444
[5]
Resource
Comments X-RAY CRYSTALLOGRAPHY (2.5 ANGSTROMS).
Medline ID 91369940
PubMed ID 1892834
Journal Biochemistry
Year 1991
Volume 30
Pages 9264-73
Authors Neidhart DJ, Howell PL, Petsko GA, Powers VM, Li RS, Kenyon GL, Gerlt JA
Title Mechanism of the reaction catalyzed by mandelate racemase. 2. Crystal structure of mandelate racemase at 2.5-A resolution: identification of the active site and possible catalytic residues.
Related PDB 2mnr
Related UniProtKB P11444
[6]
Resource
Comments
Medline ID
PubMed ID 1892833
Journal Biochemistry
Year 1991
Volume 30
Pages 9255-63
Authors Powers VM, Koo CW, Kenyon GL, Gerlt JA, Kozarich JW
Title Mechanism of the reaction catalyzed by mandelate racemase. 1. Chemical and kinetic evidence for a two-base mechanism.
Related PDB
Related UniProtKB
[7]
Resource
Comments
Medline ID
PubMed ID 1986411
Journal Science
Year 1991
Volume 251
Pages 31-2
Authors Hoffman M
Title On the road to mandelate ... racemase.
Related PDB
Related UniProtKB
[8]
Resource
Comments
Medline ID
PubMed ID 8256284
Journal Trends Biochem Sci
Year 1993
Volume 18
Pages 372-6
Authors Petsko GA, Kenyon GL, Gerlt JA, Ringe D, Kozarich JW
Title On the origin of enzymatic species.
Related PDB
Related UniProtKB
[9]
Resource
Comments X-ray crystallography
Medline ID
PubMed ID 8292591
Journal Biochemistry
Year 1994
Volume 33
Pages 635-43
Authors Landro JA, Gerlt JA, Kozarich JW, Koo CW, Shah VJ, Kenyon GL, Neidhart DJ, Fujita S, Petsko GA
Title The role of lysine 166 in the mechanism of mandelate racemase from Pseudomonas putida: mechanistic and crystallographic evidence for stereospecific alkylation by (R)-alpha-phenylglycidate.
Related PDB 1mdr 1mns
Related UniProtKB
[10]
Resource
Comments
Medline ID
PubMed ID 8009219
Journal Science
Year 1994
Volume 264
Pages 1887-90
Authors Cleland WW, Kreevoy MM
Title Low-barrier hydrogen bonds and enzymic catalysis.
Related PDB
Related UniProtKB
[11]
Resource
Comments X-RAY CRYSTALLOGRAPHY (1.85 ANGSTROMS).
Medline ID 95200898
PubMed ID 7893690
Journal Biochemistry
Year 1995
Volume 34
Pages 2788-97
Authors Kallarakal AT, Mitra B, Kozarich JW, Gerlt JA, Clifton JG, Petsko GA, Kenyon GL
Title Mechanism of the reaction catalyzed by mandelate racemase: structure and mechanistic properties of the K166R mutant.
Related PDB 1mdl
Related UniProtKB P11444
[12]
Resource
Comments X-RAY CRYSTALLOGRAPHY (2.1 ANGSTROMS).
Medline ID 95200897
PubMed ID 7893689
Journal Biochemistry
Year 1995
Volume 34
Pages 2777-87
Authors Mitra B, Kallarakal AT, Kozarich JW, Gerlt JA, Clifton JG, Petsko GA, Kenyon GL
Title Mechanism of the reaction catalyzed by mandelate racemase: importance of electrophilic catalysis by glutamic acid 317.
Related PDB 1dtn
Related UniProtKB P11444
[13]
Resource
Comments
Medline ID
PubMed ID 8987982
Journal Biochemistry
Year 1996
Volume 35
Pages 16489-501
Authors Babbitt PC, Hasson MS, Wedekind JE, Palmer DR, Barrett WC, Reed GH, Rayment I, Ringe D, Kenyon GL, Gerlt JA
Title The enolase superfamily: a general strategy for enzyme-catalyzed abstraction of the alpha-protons of carboxylic acids.
Related PDB
Related UniProtKB
[14]
Resource
Comments X-ray crystallography
Medline ID
PubMed ID 8639525
Journal Biochemistry
Year 1996
Volume 35
Pages 5662-9
Authors Schafer SL, Barrett WC, Kallarakal AT, Mitra B, Kozarich JW, Gerlt JA, Clifton JG, Petsko GA, Kenyon GL
Title Mechanism of the reaction catalyzed by mandelate racemase: structure and mechanistic properties of the D270N mutant.
Related PDB 1mra
Related UniProtKB
[15]
Resource
Comments
Medline ID
PubMed ID 9048548
Journal Biochemistry
Year 1997
Volume 36
Pages 1646-56
Authors Bearne SL, Wolfenden R
Title Mandelate racemase in pieces: effective concentrations of enzyme functional groups in the transition state.
Related PDB
Related UniProtKB
[16]
Resource
Comments
Medline ID
PubMed ID 9772161
Journal Biochemistry
Year 1998
Volume 37
Pages 14358-68
Authors Gulick AM, Palmer DR, Babbitt PC, Gerlt JA, Rayment I
Title Evolution of enzymatic activities in the enolase superfamily: crystal structure of (D)-glucarate dehydratase from Pseudomonas putida.
Related PDB
Related UniProtKB
[17]
Resource
Comments
Medline ID
PubMed ID 9748211
Journal J Biol Chem
Year 1998
Volume 273
Pages 25529-32
Authors Cleland WW, Frey PA, Gerlt JA
Title The low barrier hydrogen bond in enzymatic catalysis.
Related PDB
Related UniProtKB
[18]
Resource
Comments
Medline ID
PubMed ID 9724714
Journal Proc Natl Acad Sci U S A
Year 1998
Volume 95
Pages 10396-401
Authors Hasson MS, Schlichting I, Moulai J, Taylor K, Barrett W, Kenyon GL, Babbitt PC, Gerlt JA, Petsko GA, Ringe D
Title Evolution of an enzyme active site: the structure of a new crystal form of muconate lactonizing enzyme compared with mandelate racemase and enolase.
Related PDB
Related UniProtKB
[19]
Resource
Comments
Medline ID
PubMed ID 11063568
Journal Biochemistry
Year 2000
Volume 39
Pages 13324-35
Authors St Maurice M, Bearne SL
Title Reaction intermediate analogues for mandelate racemase: interaction between Asn 197 and the alpha-hydroxyl of the substrate promotes catalysis.
Related PDB
Related UniProtKB
[20]
Resource
Comments
Medline ID
PubMed ID 11900548
Journal Biochemistry
Year 2002
Volume 41
Pages 4048-58
Authors St Maurice M, Bearne SL
Title Kinetics and thermodynamics of mandelate racemase catalysis.
Related PDB
Related UniProtKB
[21]
Resource
Comments
Medline ID
PubMed ID 12781191
Journal Bioorg Med Chem Lett
Year 2003
Volume 13
Pages 2041-4
Authors St Maurice M, Bearne SL, Lu W, Taylor SD
Title Inhibition of mandelate racemase by alpha-fluorobenzylphosphonates.
Related PDB
Related UniProtKB
[22]
Resource
Comments
Medline ID
PubMed ID 15023082
Journal Acc Chem Res
Year 2004
Volume 37
Pages 149-58
Authors Wise EL, Rayment I
Title Understanding the importance of protein structure to nature's routes for divergent evolution in TIM barrel enzymes.
Related PDB
Related UniProtKB
[23]
Resource
Comments
Medline ID
PubMed ID 14992589
Journal Biochemistry
Year 2004
Volume 43
Pages 2524-32
Authors St Maurice M, Bearne SL
Title Hydrophobic nature of the active site of mandelate racemase.
Related PDB
Related UniProtKB
[24]
Resource
Comments
Medline ID
PubMed ID 15697231
Journal Biochemistry
Year 2005
Volume 44
Pages 2059-71
Authors Kalyanaraman C, Bernacki K, Jacobson MP
Title Virtual screening against highly charged active sites: identifying substrates of alpha-beta barrel enzymes.
Related PDB
Related UniProtKB

Comments
This enzyme catalyzes the following reactions, to produce (R)-mandelate from (S)-mandelate:
(A) Isomerization; Shift of double-bond position (from O=C-C to O-C=C), forming an enolic intermediate.
(B) Isomerization; Shift of double-bond position (from C=C-O to C-C=O).
According to the literature [11], [12], [14], [18] & [22], the catalytic reaction proceeds as follows:
(A) Isomerization; Shift of double-bond position (from O=C-C to O-C=C), forming an enolic intermediate.
(A1) A carboxylate oxygen and hydroxyl oxygen are coordinated to the cofactor, magnesium ion. (This ion must neutralize the negative charge on the carboxylate oxygen atom, along with Lys164.)
(A2) Lys166 acts as (S)-specific base to deprotonate the alpha-proton (from the single-bonded carbon) of the substrate, whereas glu317 acts as a general acid to protonate the carboxylate (double-bonded) oxygen, forming an enolic intermediate.
(A3) Here, Glu317 must stabilize the intermediate, by neutralizing the negative charge on the oxygen atom.
(B) Isomerization; Shift of double-bond position (from C=C-O to C-C=O).
(B1) The enol oxygen and hydroxyl oxygen are coordinated to the cofactor, magnesium ion. (This ion must neutralize the negative charge on the enol oxygen atom, along with Lys164.) Glu317 also stabilizes the other enol oxygen.
(B2) Asp270 modulates the activity of His297, as a pKa modulator.
(B3) His297 acts as (R)-specific acid to protonate the (double-bonded) carbon atom of the enol intermediate, whereas Glu317 acts as a general base to deprotonate the enol (single-bonded) oxygen. These completes the reaction.

Created Updated
2005-05-11 2010-08-06