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Literature summary for 3.4.17.22 extracted from

  • Garcia-Pardo, J.; Tanco, S.; Diaz, L.; Dasgupta, S.; Fernandez-Recio, J.; Lorenzo, J.; Aviles, F.X.; Fricker, L.D.
    Substrate specificity of human metallocarboxypeptidase D Comparison of the two active carboxypeptidase domains (2017), PLoS ONE, 12, e0187778 .
    View publication on PubMedView publication on EuropePMC
Search for more literature for 3.4.17.22

Cloned(Commentary)

Cloned (Comment) Organism
human carboxypeptidase D (residues 32-1298) is cloned into the pTriExTM-7 expression vector that encodes an IgM secretion signal sequence and an N-terminal Strep-Tag II fusion protein. The CPD-pTrieX-7 construct ias transfected into mammalian HEK293F cells for transient expression, using polyethylenimine as transfection reagent Homo sapiens
transfection of HEK-293F cell Homo sapiens

Crystallization (Commentary)

Crystallization (Comment) Organism
comparative modeling of the active sites of CPD domains I, II and III and molecular docking of peptide substrates Homo sapiens

Protein Variants

Protein Variants Comment Organism
E350Q critical active site residue in domain I, displays more than 50% activity from pH 5.0-7.5, similar to wild-type Homo sapiens
E350Q pH optimum of mutant enzyme is identical to wild-type enzyme, pH-range with 50% of maximal activity is pH 5.0-7.5 compared to pH 5.0-7.0 for wild-type enzyme Homo sapiens
E350Q/E762Q completely inactive towards all substrates and at all pH values tested Homo sapiens
E350Q/E762Q no detectable enzyme activity at any of the pH values examined Homo sapiens
E762Q critical active site residue in domain II, displays more than 50% activity from pH 6.5 to 7.5 Homo sapiens
E762Q pH optimum of mutant enzyme is pH 7.0, compared to pH 6.5 for wild-type enzyme. pH-range with 50% of maximal activity is pH 6.5-7.5 compared to pH 5.0-7.0 for wild-type enzyme Homo sapiens
additional information enzyme with mutations in both domains I and II is completely inactive towards all substrates and at all pH values Homo sapiens

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.153
-
 dansyl-Phe-Ala-Arg pH 6.5, 37°C, wild-type enzyme Homo sapiens
0.153
-
 dansyl-Phe-Ala-Arg wild-type, pH 6.5, 37°C Homo sapiens
0.319
-
 dansyl-Phe-Ala-Arg mutant E762Q, pH 6.5, 37°C Homo sapiens
0.319
-
 dansyl-Phe-Ala-Arg pH 6.5, 37°C, mutant enzyme E762Q Homo sapiens
0.844
-
 dansyl-Phe-Ala-Arg mutant E350Q, pH 6.5, 37°C Homo sapiens
0.844
-
 dansyl-Phe-Ala-Arg pH 6.5, 37°C, mutant enzyme E350Q Homo sapiens

Localization

Localization Comment Organism GeneOntology No. Textmining
Golgi apparatus metallocarboxypeptidase D is a membrane-bound component of the trans-Golgi network that cycles to the cell surface through exocytic and endocytic pathways Homo sapiens 5794
-
membrane metallocarboxypeptidase D is a membrane-bound component of the trans-Golgi network that cycles to the cell surface through exocytic and endocytic pathways Homo sapiens 16020
-

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
170000
-
-
 Homo sapiens

Organism

Organism UniProt Comment Textmining
Homo sapiens O75976
-
-

Purification (Commentary)

Purification (Comment) Organism
-
 Homo sapiens
recombinant protein Homo sapiens

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
Dansyl-Phe-Ala-Arg + H2O
-
 Homo sapiens Dansyl-Phe-Ala + Arg
-
 ?
Dansyl-Phe-Gly-Arg + H2O
-
 Homo sapiens Dansyl-Phe-Gly + Arg
-
 ?
additional information CPD cleaves C-terminal Lys or Arg from a subset of the peptides. Most of the identified substrates of domain I contain C-terminal Arg, whereas comparable numbers of Lys- and Arg-containing peptides are substrates of domain II. Some peptides with C-terminal basic residues are not cleaved by either domain I or II Homo sapiens ?
-
 ?
additional information multicatalytic enzyme with three carboxypeptidase-like domains, although only the first two domains are predicted to be enzymatically active. The enzyme cleaves exclusively C-terminal basic residues. A quantitative peptidomics approach is used to compare the activities of CPD domains I and II towards a large number of peptides. the enzyme cleaves C-terminal Lys or Arg from a subset of the peptides. Most of the identified substrates of domain I contain C-terminal Arg, whereas comparable numbers of Lys- and Arg-containing peptides are substrates of domain II. Some peptides with C-terminal basic residues are not cleaved by either domain I or II, showing the importance of the P1 position for CPD activity. The preference of domain I for C-terminal Arg is validated through molecular docking experiments Homo sapiens ?
-
 ?
additional information no substrate: dansyl-Phe-Pro-Arg Homo sapiens ?
-
 ?

Synonyms

Synonyms Comment Organism
CPD
-
 Homo sapiens

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
7
-
 dansyl-Phe-Ala-Arg mutant E350Q, pH 6.5, 37°C Homo sapiens
7
-
 dansyl-Phe-Ala-Arg pH 6.5, 37°C, mutant enzyme E350Q Homo sapiens
8.5
-
 dansyl-Phe-Ala-Arg mutant E762Q, pH 6.5, 37°C Homo sapiens
8.5
-
 dansyl-Phe-Ala-Arg pH 6.5, 37°C, mutant enzyme E762Q Homo sapiens
12.5
-
 dansyl-Phe-Ala-Arg pH 6.5, 37°C, wild-type enzyme Homo sapiens
12.5
-
 dansyl-Phe-Ala-Arg wild-type, pH 6.5, 37°C Homo sapiens

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
6.5
-
-
 Homo sapiens
6.5
-
 enzyme with all three domains intact and mutant enzyme E350Q Homo sapiens
7
-
 mutant enzyme E762Q Homo sapiens

pH Range

pH Minimum pH Maximum Comment Organism
5 7 enzyme with all three domains intact and enzyme with mutation of domain I display more than 50% activity from pH 5.0 to 7.5 Homo sapiens
5 7.5 mutant enzyme E350Q displays more than 50% activity from pH 5.0 to 7.5 Homo sapiens
5 7.5 wild-type, more than 50% of maximum activity within Homo sapiens
6.5 7.5 mutant enzyme E762Q displays more than 50% activity from pH 6.5 to 7.5 Homo sapiens

General Information

General Information Comment Organism
physiological function together with the differences in pH optima, the different substrate specificities of CPD domains I and II allow the enzyme to perform distinct functions in the various locations within the cell Homo sapiens

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
8
-
 dansyl-Phe-Ala-Arg mutant E350Q, pH 6.5, 37°C Homo sapiens
8.3
-
 dansyl-Phe-Ala-Arg pH 6.5, 37°C, mutant enzyme E350Q Homo sapiens
26.6
-
 dansyl-Phe-Ala-Arg pH 6.5, 37°C, mutant enzyme E762Q Homo sapiens
27
-
 dansyl-Phe-Ala-Arg mutant E762Q, pH 6.5, 37°C Homo sapiens
81.7
-
 dansyl-Phe-Ala-Arg pH 6.5, 37°C, wild-type enzyme Homo sapiens
82
-
 dansyl-Phe-Ala-Arg wild-type, pH 6.5, 37°C Homo sapiens