Cloned (Comment) | Organism |
---|---|
N-terminal His-tagged SySSADH expressed in Escherichia coli B834 (DE3) methionine auxotroph cells | Synechococcus sp. |
Crystallization (Comment) | Organism |
---|---|
crystal structures of SySSADH determined in their apo form, as a binary complex with NADP+ and as a ternary complex with succinic semialdehyde and NADPH, resoultion of 1.7 A for the apo form and of 1.4 A for the binary and ternary complex | Synechococcus sp. |
Protein Variants | Comment | Organism |
---|---|---|
C262A | active site mutation, nonfunctional because Cys-262 acts as a nucleophilel | Synechococcus sp. |
E228A | active site mutation, nonfunctional because Glu-228 acts as a general base | Synechococcus sp. |
E228Q | active site mutation, nonfunctional because Glu-228 acts as a general base | Synechococcus sp. |
F132A | activity of about 1030% of the wild type enzyme, indicating a contribution of these succinic semialdehyde binding residues to the overall enzyme activity | Synechococcus sp. |
F425A | inactive, suggesting that Phe-425 plays an important role in substrate binding | Synechococcus sp. |
I263A | activity of about 1030% of the wild type enzyme, indicating a contribution of these succinic semialdehyde binding residues to the overall enzyme activity | Synechococcus sp. |
N131A | mutation of a residue that interacts with the O4 atom or the carboxyl group of succinic semialdehyde thus abolishing enzyme activity | Synechococcus sp. |
N131D | mutation of a residue that interacts with the O4 atom or the carboxyl group of succinic semialdehyde thus abolishing enzyme activity | Synechococcus sp. |
R139A | activity of about 1030% of the wild type enzyme, indicating a contribution of these succinic semialdehyde binding residues to the overall enzyme activity | Synechococcus sp. |
R139K | mutant enzyme exhibited an activity up to 80% that of the wild type enzyme, suggesting the significance of a positively charged residue in the binding of the carboxyl group of succinic semialdehyde | Synechococcus sp. |
S419A | mutation of a residue that interacts with the O4 atom or the carboxyl group of succinic semialdehyde thus abolishing enzyme activity | Synechococcus sp. |
W135A | activity of about 1030% of the wild type enzyme, indicating a contribution of these succinic semialdehyde binding residues to the overall enzyme activity | Synechococcus sp. |
Inhibitors | Comment | Organism | Structure |
---|---|---|---|
H2O2 | 50 microM H2O2 sharply reduces activity to 31% of the H2O2-free enzyme and activity further decreases to 3% at 1 mM H2O2 | Synechococcus sp. |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
additional information | - |
additional information | the Km-value of succinate semialdehyde estimated to be far less than 0.05 mM | Synechococcus sp. | |
0.439 | - |
NADP+ | 30°C, pH 7.6 | Synechococcus sp. |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
succinate semialdehyde + NADP+ + H2O | Synechococcus sp. | data from crystal structures provide details about the catalytic mechanism by revealing a covalent adduct of a cofactor with the catalytic cysteine in the binary complex and a proposed thiohemiacetal intermediate in the ternary complex | succinate + NADPH + 2 H+ | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Synechococcus sp. | B1XMM6 | - |
- |
Oxidation Stability | Organism |
---|---|
SySSADH is an oxidation-sensitive enzyme and the formation of the NADP-cysteine adduct is a kinetically preferred event that protects the catalytic Cys-262 from H2O2-dependent oxidative stress | Synechococcus sp. |
Purification (Comment) | Organism |
---|---|
using immobilized metal affinity chromatography, removal of N-terminal His tag, further purification by immobilized metal affinity chromatography and gel filtration using a Superdex 200 column | Synechococcus sp. |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
succinate semialdehyde + NADP+ + H2O | data from crystal structures provide details about the catalytic mechanism by revealing a covalent adduct of a cofactor with the catalytic cysteine in the binary complex and a proposed thiohemiacetal intermediate in the ternary complex | Synechococcus sp. | succinate + NADPH + 2 H+ | - |
? |
Subunits | Comment | Organism |
---|---|---|
homodimer | gel filtration, SySSADH monomer consists of three segments - alpha/beta-fold N- and C-domains for a cofactor binding and a catalytic domain, and three antiparallel beta-strands constituting a dimerization domain | Synechococcus sp. |
Synonyms | Comment | Organism |
---|---|---|
SSADH | - |
Synechococcus sp. |
SySSADH | - |
Synechococcus sp. |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
5 | - |
NADP+ | 30°C, pH 7.6 | Synechococcus sp. |
General Information | Comment | Organism |
---|---|---|
metabolism | succinic semialdehyde dehydrogenase from Synechococcus is an essential enzyme in the tricarboxylic acid cycle of cyanobacteria | Synechococcus sp. |