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Literature summary for 6.3.1.19 extracted from

  • Regev, O.; Roth, Z.; Korman, M.; Khalaila, I.; Gur, E.
    A kinetic model for the prevalence of mono- over poly-pupylation (2015), FEBS J., 282, 4176-4186 .
    View publication on PubMed
Search for more literature for 6.3.1.19

Cloned(Commentary)

Cloned (Comment) Organism
recombinant expression of His6-tagged enzyme in Escherichia coli strain ER2566 Mycolicibacterium smegmatis

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
 additional information Michaelis-Menten kinetic analysis of poly-pupylation, overview Mycolicibacterium smegmatis
0.22
-
 [IdeR]-L-lysine pH 7.5, 30°C, recombinant His-tagged enzyme Mycolicibacterium smegmatis

Metals/Ions

Metals/Ions Comment Organism Structure
Mg2+ required Mycolicibacterium smegmatis

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
ATP + [prokaryotic ubiquitin-like protein]-L-glutamate + [protein]-L-lysine Mycolicibacterium smegmatis
-
 ADP + phosphate + N6-([prokaryotic ubiquitin-like protein]-gamma-L-glutamyl)-[protein]-L-lysine
-
 ?
ATP + [prokaryotic ubiquitin-like protein]-L-glutamate + [protein]-L-lysine Mycolicibacterium smegmatis ATCC 700084 / mc2155
-
 ADP + phosphate + N6-([prokaryotic ubiquitin-like protein]-gamma-L-glutamyl)-[protein]-L-lysine
-
 ?
additional information Mycolicibacterium smegmatis PafA can pupylate itself, extra-pupylation by PafA can also occur, mostly in the form of poly-pupylation. Unlike in the eukaryotic UPS where poly-ubiquitylation is seen, in vivo targets of the Pup-proteasome system are found almost exclusively to be monopupylated, except for PafA itself. Poly-pupylation occurs via pupylation of an already pupylated target, rather than by conjugation of pre-formed poly-Pup chains ?
-
 ?
additional information Mycolicibacterium smegmatis ATCC 700084 / mc2155 PafA can pupylate itself, extra-pupylation by PafA can also occur, mostly in the form of poly-pupylation. Unlike in the eukaryotic UPS where poly-ubiquitylation is seen, in vivo targets of the Pup-proteasome system are found almost exclusively to be monopupylated, except for PafA itself. Poly-pupylation occurs via pupylation of an already pupylated target, rather than by conjugation of pre-formed poly-Pup chains ?
-
 ?

Organism

Organism UniProt Comment Textmining
Mycolicibacterium smegmatis A0QZ42
-
-
Mycolicibacterium smegmatis ATCC 700084 / mc2155 A0QZ42
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant His6-tagged enzyme from Escherichia coli strain ER2566 by nickel affinity chormatography Mycolicibacterium smegmatis

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
ATP + [prokaryotic ubiquitin-like protein]-L-glutamate + [IdeR]-L-lysine Mycobacterium smegmatis recombinant His6-tagged IdeR is a model substrate for in vitro studies Mycolicibacterium smegmatis ADP + phosphate + N6-([prokaryotic ubiquitin-like protein]-gamma-L-glutamyl)-[IdeR]-L-lysine
-
 ?
ATP + [prokaryotic ubiquitin-like protein]-L-glutamate + [IdeR]-L-lysine Mycobacterium smegmatis recombinant His6-tagged IdeR is a model substrate for in vitro studies Mycolicibacterium smegmatis ATCC 700084 / mc2155 ADP + phosphate + N6-([prokaryotic ubiquitin-like protein]-gamma-L-glutamyl)-[IdeR]-L-lysine
-
 ?
ATP + [prokaryotic ubiquitin-like protein]-L-glutamate + [PanB]-L-lysine PanB is a model substrate of PafA Mycolicibacterium smegmatis ADP + phosphate + N6-([prokaryotic ubiquitin-like protein]-gamma-L-glutamyl)-[PanB]-L-lysine
-
 ?
ATP + [prokaryotic ubiquitin-like protein]-L-glutamate + [prokaryotic ubiquitin-like protein]-L-lysine in addition to the lysine at position 61, Pup (prokaryotic ubiquitin-like protein) presents two more lysines, one at position 7 and another at position 31. Pup can be pupylated on different lysines Mycolicibacterium smegmatis ADP + phosphate + N6-([prokaryotic ubiquitin-like protein]-gamma-L-glutamyl)-[prokaryotic ubiquitin-like protein]-L-lysine
-
 ?
ATP + [prokaryotic ubiquitin-like protein]-L-glutamate + [protein]-L-lysine
-
 Mycolicibacterium smegmatis ADP + phosphate + N6-([prokaryotic ubiquitin-like protein]-gamma-L-glutamyl)-[protein]-L-lysine
-
 ?
ATP + [prokaryotic ubiquitin-like protein]-L-glutamate + [protein]-L-lysine the Pup variant used presents a glutamate at its C-terminus (PupE) and, as such, can be readily conjugated to target proteins by PafA Mycolicibacterium smegmatis ADP + phosphate + N6-([prokaryotic ubiquitin-like protein]-gamma-L-glutamyl)-[protein]-L-lysine
-
 ?
ATP + [prokaryotic ubiquitin-like protein]-L-glutamate + [protein]-L-lysine
-
 Mycolicibacterium smegmatis ATCC 700084 / mc2155 ADP + phosphate + N6-([prokaryotic ubiquitin-like protein]-gamma-L-glutamyl)-[protein]-L-lysine
-
 ?
ATP + [prokaryotic ubiquitin-like protein]-L-glutamate + [protein]-L-lysine the Pup variant used presents a glutamate at its C-terminus (PupE) and, as such, can be readily conjugated to target proteins by PafA Mycolicibacterium smegmatis ATCC 700084 / mc2155 ADP + phosphate + N6-([prokaryotic ubiquitin-like protein]-gamma-L-glutamyl)-[protein]-L-lysine
-
 ?
additional information PafA can pupylate itself, extra-pupylation by PafA can also occur, mostly in the form of poly-pupylation. Unlike in the eukaryotic UPS where poly-ubiquitylation is seen, in vivo targets of the Pup-proteasome system are found almost exclusively to be monopupylated, except for PafA itself. Poly-pupylation occurs via pupylation of an already pupylated target, rather than by conjugation of pre-formed poly-Pup chains Mycolicibacterium smegmatis ?
-
 ?
additional information PafA binds IdeR with low affinity, compared to PanB, a model substrate whose affinity for PafA is about 10fold higher Mycolicibacterium smegmatis ?
-
 ?
additional information PafA can pupylate itself, extra-pupylation by PafA can also occur, mostly in the form of poly-pupylation. Unlike in the eukaryotic UPS where poly-ubiquitylation is seen, in vivo targets of the Pup-proteasome system are found almost exclusively to be monopupylated, except for PafA itself. Poly-pupylation occurs via pupylation of an already pupylated target, rather than by conjugation of pre-formed poly-Pup chains Mycolicibacterium smegmatis ATCC 700084 / mc2155 ?
-
 ?
additional information PafA binds IdeR with low affinity, compared to PanB, a model substrate whose affinity for PafA is about 10fold higher Mycolicibacterium smegmatis ATCC 700084 / mc2155 ?
-
 ?

Subunits

Subunits Comment Organism
? x * 51000, recombinant His6-tagged enzyme, SDS-PAGE Mycolicibacterium smegmatis

Synonyms

Synonyms Comment Organism
PafA
-
 Mycolicibacterium smegmatis
Pup ligase
-
 Mycolicibacterium smegmatis

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
30
-
 assay at Mycolicibacterium smegmatis

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.014
-
 [IdeR]-L-lysine pH 7.5, 30°C, recombinant His-tagged enzyme Mycolicibacterium smegmatis

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.5
-
 assay at Mycolicibacterium smegmatis

Cofactor

Cofactor Comment Organism Structure
ATP
-
 Mycolicibacterium smegmatis

General Information

General Information Comment Organism
physiological function prokaryotic ubiquitin-like protein, Pup, is conjugated to proteins by PafA, the only Pup ligase identified thus far, through the formation of an iso-peptide bond between the gamma-carboxylate of a glutamate side chain at the C terminus of Pup and the epsilon-amine of a lysine residue on the target protein. Pupylation is a cytoplasmic signal for proteasomal degradation. Pup ligase PafA conjugates the small protein Pup to lysine side chains of target proteins. Mono-Pup moieties are almost exclusively observed in vivo and are sufficient as degradation tags Mycolicibacterium smegmatis