Crystallization (Comment) | Organism |
---|---|
mutant D154A in complex with MgATP-RNA, hanging drop vapour diffusion method, in 0.1 M bis-Tris propane, pH 6.4, 0.2 M Li-acetate, and 16% PEG 3350 | Saccharomyces cerevisiae |
mutant D154A, trapped in complex with ATP and a five residue poly(A). Enzyme has undergone significant domain movement and shows a closed conformation with extensive interactions between substrates and all three polymerase domains | Saccharomyces cerevisiae |
Protein Variants | Comment | Organism |
---|---|---|
D154A | crystallization data, closed conformation with extensive interactions between substrates and all three polymerase domains | Saccharomyces cerevisiae |
D154A | the mutant has a nearly identical melting temperature as wild type PAP | Saccharomyces cerevisiae |
K215A | 2- to 4fold increase in Km values | Saccharomyces cerevisiae |
K215A | the mutant has a nearly identical melting temperature as wild type PAP | Saccharomyces cerevisiae |
N189A | residue bridges the N and middle domains in the closed state | Saccharomyces cerevisiae |
N189A | the mutant has a nearly identical melting temperature as wild type PAP | Saccharomyces cerevisiae |
N226A | mutation affects the equilibrium between the open- and closed-domain forms of the enzyme | Saccharomyces cerevisiae |
N226A | the mutant has a nearly identical melting temperature as wild type PAP | Saccharomyces cerevisiae |
Y224F | mutation affects the equilibrium between the open- and closed-domain forms of the enzyme | Saccharomyces cerevisiae |
Y224F | altered melting temperature (44°C) compared to the wild type enzyme | Saccharomyces cerevisiae |
General Stability | Organism |
---|---|
substrates poly(A) and Mg-ATP induce the conformational change, resulting in stabilization of the closed enzyme state and enabling catalysis | Saccharomyces cerevisiae |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.036 | - |
ATP | wild-type, pH 7.0, 30°C | Saccharomyces cerevisiae | |
0.0468 | - |
(A)n | wild-type, pH 7.0, 30°C | Saccharomyces cerevisiae | |
0.055 | - |
GTP | mutant N226A, pH 7.0, 30°C | Saccharomyces cerevisiae | |
0.062 | - |
GTP | wild-type, pH 7.0, 30°C | Saccharomyces cerevisiae | |
0.08 | - |
ATP | mutant N189A, pH 7.0, 30°C | Saccharomyces cerevisiae | |
0.104 | - |
CTP | wild-type, pH 7.0, 30°C | Saccharomyces cerevisiae | |
0.106 | - |
(A)n | mutant N189A, pH 7.0, 30°C | Saccharomyces cerevisiae | |
0.14 | - |
CTP | mutant N226A, pH 7.0, 30°C | Saccharomyces cerevisiae | |
0.148 | - |
CTP | mutant N189A, pH 7.0, 30°C | Saccharomyces cerevisiae | |
0.195 | - |
(A)n | mutant K215A, pH 7.0, 30°C | Saccharomyces cerevisiae | |
0.249 | - |
ATP | mutant N226A, pH 7.0, 30°C | Saccharomyces cerevisiae | |
0.367 | - |
(A)n | mutant N226A, pH 7.0, 30°C | Saccharomyces cerevisiae | |
0.368 | - |
CTP | mutant K215A, pH 7.0, 30°C | Saccharomyces cerevisiae | |
0.406 | - |
ATP | mutant K215A, pH 7.0, 30°C | Saccharomyces cerevisiae | |
0.711 | - |
(A)n | mutant Y224F, pH 7.0, 30°C | Saccharomyces cerevisiae | |
0.929 | - |
ATP | mutant Y224F, pH 7.0, 30°C | Saccharomyces cerevisiae | |
4.7 | - |
CTP | mutant Y224F, pH 7.0, 30°C | Saccharomyces cerevisiae |
Metals/Ions | Comment | Organism | Structure |
---|---|---|---|
Mg2+ | essential for activity | Saccharomyces cerevisiae |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Saccharomyces cerevisiae | P29468 | - |
- |
Purification (Comment) | Organism |
---|---|
nickel affinity column chromatography and ion exchange chromatography | Saccharomyces cerevisiae |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
(A)n + ATP | - |
Saccharomyces cerevisiae | (A)n+1 + diphosphate | - |
? | |
(A)n + CTP | - |
Saccharomyces cerevisiae | (A)n-C + diphosphate | - |
? | |
(A)n + diphosphate | - |
Saccharomyces cerevisiae | (A)n-1 + ATP | - |
? | |
(A)n + GTP | - |
Saccharomyces cerevisiae | (A)n-G + diphosphate | - |
? | |
ATP + RNA | - |
Saccharomyces cerevisiae | diphosphate + RNA(A)n | - |
? |
Synonyms | Comment | Organism |
---|---|---|
PAP | PAP is a template-independent polymerase that belongs to the DNA polymerase beta family of enzymes | Saccharomyces cerevisiae |
poly(A) polymerase | - |
Saccharomyces cerevisiae |
Temperature Stability Minimum [°C] | Temperature Stability Maximum [°C] | Comment | Organism |
---|---|---|---|
30 | 48 | remains stable at 30°C, the melting temperature for the wild type enzyme is at 48°C | Saccharomyces cerevisiae |