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Literature summary extracted from

  • Al-Dabbagh, B.; Henry, X.; El Ghachi, M.; Auger, G.; Blanot, D.; Parquet, C.; Mengin-Lecreulx, D.; Bouhss, A.
    Active site mapping of MraY, a member of the polyprenyl-phosphate N-acetylhexosamine 1-phosphate transferase superfamily, catalyzing the first membrane step of peptidoglycan biosynthesis (2008), Biochemistry, 47, 8919-8928.
    View publication on PubMed

Application

EC Number Application Comment Organism
2.7.8.13 drug development antibacterial chemotherapy against pathogenic bacteria being multiresistant to common antibiotics Bacillus subtilis

Cloned(Commentary)

EC Number Cloned (Comment) Organism
2.7.8.13 in pET28b for site-directed mutagenesis and expression with N-terminal hexa-His-tag in Escherichia coli C43(DE3) or functional complementation in BW25113-derived thermosensitive strain (EcoliTsMraY) lacking endogenous MraY Bacillus subtilis

Protein Variants

EC Number Protein Variants Comment Organism
2.7.8.13 D174N 0.39% of wild-type activity, 70% of optimum activity in presence of 250 mM Mg2+, altered catalytic efficiency (kcat), not capable of functional complementation, invariant residue in third cytoplasmic segment Bacillus subtilis
2.7.8.13 D177N 1.6% of wild-type activity, 70% of optimum activity in presence of 250 mM Mg2+, altered catalytic efficiency (kcat), invariant residue in third cytoplasmic segment Bacillus subtilis
2.7.8.13 D231N 0.42% of wild-type activity, altered catalytic efficiency (kcat), not capable of functional complementation, invariant residue in fourth cytoplasmic segment Bacillus subtilis
2.7.8.13 D98N 0.014% of wild-type activity, shifted pH optimum (pH 9.0-9.4) compared to wild-type (pH 7.5), highly decreased turnover (kcat, pH 7.2, elevated turnover at pH 9.4), decreased KM for UDP-N-acetylmuramoyl-pentapeptide (pH 7.2 and pH 9.4), increased KM for undecaprenyl phosphate (2fold at pH 7.2, 6fold at pH 9.4), not capable of functional complementation, invariant residue in second cytoplasmic segment Bacillus subtilis
2.7.8.13 D99N 0.0076% of wild-type activity, highly decreased catalytic efficiency (kcat), increased KM for undecaprenyl phosphate, not capable of functional complementation, invariant residue in second cytoplasmic segment Bacillus subtilis
2.7.8.13 E299Q 11% of wild-type activity, catalytic activity similarly affected by the presence of 250 mM Mg2+ as wild-type, slightly altered catalytic efficiency (kcat, 10%), invariant residue in fifth cytoplasmic segment Bacillus subtilis
2.7.8.13 H289R 0.0012% of wild-type activity, highly decreased catalytic efficiency (kcat), not capable of functional complementation, invariant residue in fifth cytoplasmic segment Bacillus subtilis
2.7.8.13 H290R 0.14% of wild-type activity, altered catalytic efficiency (kcat), not capable of functional complementation, invariant residue in fifth cytoplasmic segment Bacillus subtilis
2.7.8.13 H291R 0.50% of wild-type activity, altered catalytic efficiency (kcat), increased KM for UDP-N-acetylmuramoyl-pentapeptide, invariant residue in fifth cytoplasmic segment Bacillus subtilis
2.7.8.13 H45R 0.54% of wild-type activity, 70% of optimum activity in presence of 250 mM Mg2+, altered catalytic efficiency (kcat), increased KM for UDP-N-acetylmuramoyl-pentapeptide, invariant residue in first cytoplasmic segment Bacillus subtilis
2.7.8.13 K116Q 0.046% of wild-type activity, highly decreased catalytic efficiency (kcat), not capable of functional complementation, invariant residue in second cytoplasmic segment Bacillus subtilis
2.7.8.13 K48Q 0.51% of wild-type activity, altered catalytic efficiency (kcat), increased KM for UDP-N-acetylmuramoyl-pentapeptide, invariant residue in first cytoplasmic segment Bacillus subtilis
2.7.8.13 N168A 1.5% of wild-type activity, altered catalytic efficiency (kcat), invariant residue in third cytoplasmic segment Bacillus subtilis
2.7.8.13 N171A 0.34% of wild-type activity, altered catalytic efficiency (kcat), not capable of functional complementation, invariant residue in third cytoplasmic segment Bacillus subtilis

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
2.7.8.13 additional information
-
 undecaprenyl phosphate 3fold increase in KM for mutant D98N at pH 9.4 (ca. 1.2 mM) compared to pH 7.2 Bacillus subtilis
2.7.8.13 additional information
-
 UDP-N-acetylmuramoyl-pentapeptide no increase in KM for mutant D98N at pH 9.4 compared to pH 7.2 Bacillus subtilis
2.7.8.13 0.15
-
 undecaprenyl phosphate mutant H290R Bacillus subtilis
2.7.8.13 0.16
-
 undecaprenyl phosphate wild-type Bacillus subtilis
2.7.8.13 0.17
-
 undecaprenyl phosphate mutant D174N Bacillus subtilis
2.7.8.13 0.17
-
 undecaprenyl phosphate mutant D231N Bacillus subtilis
2.7.8.13 0.21
-
 undecaprenyl phosphate mutant N168A Bacillus subtilis
2.7.8.13 0.22
-
 undecaprenyl phosphate mutant E299Q Bacillus subtilis
2.7.8.13 0.22
-
 undecaprenyl phosphate mutant H45R Bacillus subtilis
2.7.8.13 0.22
-
 undecaprenyl phosphate mutant K116Q Bacillus subtilis
2.7.8.13 0.23
-
 undecaprenyl phosphate mutant D177N Bacillus subtilis
2.7.8.13 0.24
-
 undecaprenyl phosphate mutant N171A Bacillus subtilis
2.7.8.13 0.25
-
 undecaprenyl phosphate mutant H289R Bacillus subtilis
2.7.8.13 0.26
-
 undecaprenyl phosphate mutant K48Q Bacillus subtilis
2.7.8.13 0.27
-
 undecaprenyl phosphate mutant H291R Bacillus subtilis
2.7.8.13 0.32
-
 undecaprenyl phosphate mutant D98N Bacillus subtilis
2.7.8.13 0.34
-
 UDP-N-acetylmuramoyl-pentapeptide mutant D98N Bacillus subtilis
2.7.8.13 0.35
-
 undecaprenyl phosphate mutant D99N Bacillus subtilis
2.7.8.13 0.73
-
 UDP-N-acetylmuramoyl-pentapeptide mutant N171A Bacillus subtilis
2.7.8.13 0.76
-
 UDP-N-acetylmuramoyl-pentapeptide mutant D99N Bacillus subtilis
2.7.8.13 0.86
-
 UDP-N-acetylmuramoyl-pentapeptide mutant D177N Bacillus subtilis
2.7.8.13 0.94
-
 UDP-N-acetylmuramoyl-pentapeptide wild-type Bacillus subtilis
2.7.8.13 0.95
-
 UDP-N-acetylmuramoyl-pentapeptide mutant N168A Bacillus subtilis
2.7.8.13 0.99
-
 UDP-N-acetylmuramoyl-pentapeptide mutant D174N Bacillus subtilis
2.7.8.13 1
-
 UDP-N-acetylmuramoyl-pentapeptide mutant D231N Bacillus subtilis
2.7.8.13 1.1
-
 UDP-N-acetylmuramoyl-pentapeptide mutant H289R Bacillus subtilis
2.7.8.13 1.3
-
 UDP-N-acetylmuramoyl-pentapeptide mutant H290R Bacillus subtilis
2.7.8.13 1.3
-
 UDP-N-acetylmuramoyl-pentapeptide mutant K48Q Bacillus subtilis
2.7.8.13 1.4
-
 UDP-N-acetylmuramoyl-pentapeptide mutant E299Q Bacillus subtilis
2.7.8.13 2
-
 UDP-N-acetylmuramoyl-pentapeptide mutant K116Q Bacillus subtilis
2.7.8.13 3.4
-
 UDP-N-acetylmuramoyl-pentapeptide mutant H291R Bacillus subtilis
2.7.8.13 3.4
-
 UDP-N-acetylmuramoyl-pentapeptide mutant H45R Bacillus subtilis

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
2.7.8.13 Mg2+ optimum concentration for wild-type activity: 30-60 mM, inhibitory above 100 mM, 2% of optimum activity left at 250 mM for wild-type and 30-70% for mutants (except mutant E299Q) Bacillus subtilis

Organism

EC Number Organism UniProt Comment Textmining
2.7.8.13 Bacillus subtilis Q03521
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
2.7.8.13 from membrane extracts in presence of DDM detergent and by Ni-NTA metal affinity chromatography Bacillus subtilis

Specific Activity [micromol/min/mg]

EC Number Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
2.7.8.13 0.00002
-
 mutant H289R Bacillus subtilis
2.7.8.13 0.00013
-
 mutant D99N Bacillus subtilis
2.7.8.13 0.00024
-
 mutant D98N Bacillus subtilis
2.7.8.13 0.00078
-
 mutant K116Q Bacillus subtilis
2.7.8.13 0.0024
-
 mutant H290R Bacillus subtilis
2.7.8.13 0.0058
-
 mutant N171A Bacillus subtilis
2.7.8.13 0.0066
-
 mutant D174N Bacillus subtilis
2.7.8.13 0.0072
-
 mutant D231N Bacillus subtilis
2.7.8.13 0.0086
-
 mutant H291R Bacillus subtilis
2.7.8.13 0.0087
-
 mutant K48Q Bacillus subtilis
2.7.8.13 0.0092
-
 mutant H45R Bacillus subtilis
2.7.8.13 0.026
-
 mutant N168A Bacillus subtilis
2.7.8.13 0.028
-
 mutant D177N Bacillus subtilis
2.7.8.13 0.18
-
 mutant E299Q Bacillus subtilis
2.7.8.13 1.7
-
 wild-type Bacillus subtilis

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
2.7.8.13 additional information all five cytoplasmic segments contribute to catalytic site with at least 14 invariant polar and/or charged, non-clustered residues, conserved residue D98 supposedly involved in deprotonation of lipid substrate during catalysis at physiological pH 7.4 (pKa for undecaprenyl phosphate: 6.6 and 9.2) Bacillus subtilis ?
-
 ?
2.7.8.13 additional information functional complementation in a MraY-lacking, thermosensitive Escherichia coli mutant strain (2fold increased UDP-N-acetylmuramoyl-pentapeptide level compared to wild-type cells when grown 30 min at 42°C, depletion of lipid I) by overexpression of wild-type or mutants E299Q, D177N, N168A, H45R, K48Q, H291R Bacillus subtilis ?
-
 ?
2.7.8.13 UDP-N-acetylmuramoyl-pentapeptide + undecaprenyl phosphate 25°C, pH 7.2 Bacillus subtilis UMP + N-acetylmuramoyl-pentapeptide-diphosphoundecaprenol lipid I synthesis, peptidoglycan biosynthesis ?

Synonyms

EC Number Synonyms Comment Organism
2.7.8.13 MraY
-
 Bacillus subtilis
2.7.8.13 MraY transferase
-
 Bacillus subtilis

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
2.7.8.13 additional information
-
 undecaprenyl phosphate 11fold increase in kcat for mutant D98N at pH 9.4 (ca. 0.0083 1/sec) compared to pH 7.2 Bacillus subtilis
2.7.8.13 additional information
-
 UDP-N-acetylmuramoyl-pentapeptide 11fold increase in kcat for mutant D98N at pH 9.4 (ca. 0.0083 1/sec) compared to pH 7.2 Bacillus subtilis
2.7.8.13 0.00005
-
 undecaprenyl phosphate mutant H289R Bacillus subtilis
2.7.8.13 0.00005
-
 UDP-N-acetylmuramoyl-pentapeptide mutant H289R Bacillus subtilis
2.7.8.13 0.00083
-
 undecaprenyl phosphate mutant D99N Bacillus subtilis
2.7.8.13 0.00083
-
 UDP-N-acetylmuramoyl-pentapeptide mutant D99N Bacillus subtilis
2.7.8.13 0.00117
-
 undecaprenyl phosphate mutant D98N Bacillus subtilis
2.7.8.13 0.00117
-
 UDP-N-acetylmuramoyl-pentapeptide mutant D98N Bacillus subtilis
2.7.8.13 0.0015
-
 undecaprenyl phosphate mutant K116Q Bacillus subtilis
2.7.8.13 0.0015
-
 UDP-N-acetylmuramoyl-pentapeptide mutant K116Q Bacillus subtilis
2.7.8.13 0.00383
-
 undecaprenyl phosphate mutant H290R Bacillus subtilis
2.7.8.13 0.00383
-
 UDP-N-acetylmuramoyl-pentapeptide mutant H290R Bacillus subtilis
2.7.8.13 0.0123
-
 undecaprenyl phosphate mutant D174N Bacillus subtilis
2.7.8.13 0.0123
-
 UDP-N-acetylmuramoyl-pentapeptide mutant D174N Bacillus subtilis
2.7.8.13 0.0135
-
 undecaprenyl phosphate mutant N171A Bacillus subtilis
2.7.8.13 0.0135
-
 UDP-N-acetylmuramoyl-pentapeptide mutant N171A Bacillus subtilis
2.7.8.13 0.0163
-
 undecaprenyl phosphate mutant D231N Bacillus subtilis
2.7.8.13 0.0163
-
 UDP-N-acetylmuramoyl-pentapeptide mutant D231N Bacillus subtilis
2.7.8.13 0.023
-
 undecaprenyl phosphate mutant H45R Bacillus subtilis
2.7.8.13 0.023
-
 UDP-N-acetylmuramoyl-pentapeptide mutant H45R Bacillus subtilis
2.7.8.13 0.027
-
 undecaprenyl phosphate mutant K48Q Bacillus subtilis
2.7.8.13 0.027
-
 UDP-N-acetylmuramoyl-pentapeptide mutant K48Q Bacillus subtilis
2.7.8.13 0.0317
-
 undecaprenyl phosphate mutant H291R Bacillus subtilis
2.7.8.13 0.0317
-
 UDP-N-acetylmuramoyl-pentapeptide mutant H291R Bacillus subtilis
2.7.8.13 0.065
-
 undecaprenyl phosphate mutant D177N Bacillus subtilis
2.7.8.13 0.065
-
 UDP-N-acetylmuramoyl-pentapeptide mutant D177N Bacillus subtilis
2.7.8.13 0.0717
-
 undecaprenyl phosphate mutant N168A Bacillus subtilis
2.7.8.13 0.0717
-
 UDP-N-acetylmuramoyl-pentapeptide mutant N168A Bacillus subtilis
2.7.8.13 0.53
-
 undecaprenyl phosphate mutant E299Q Bacillus subtilis
2.7.8.13 0.53
-
 UDP-N-acetylmuramoyl-pentapeptide mutant E299Q Bacillus subtilis
2.7.8.13 5.3
-
 undecaprenyl phosphate wild-type Bacillus subtilis
2.7.8.13 5.3
-
 UDP-N-acetylmuramoyl-pentapeptide wild-type Bacillus subtilis

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
2.7.8.13 7.5
-
 wild-type and all mutants except mutant D98N Bacillus subtilis
2.7.8.13 9.4
-
 9.0-9.4, mutant D98N Bacillus subtilis

pH Range

EC Number pH Minimum pH Maximum Comment Organism
2.7.8.13 6.8 9.4 mutant D98N shows different pH profile (specific activity ratio between pH9.4/pH7.6: 2.5) than wild-type (and all mutants except mutant D98N, specific activity ratio between pH9.4/pH7.6: 0.22-0.83) Bacillus subtilis