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Literature summary for 1.2.1.79 extracted from

  • Phonbuppha, J.; Maenpuen, S.; Munkajohnpong, P.; Chaiyen, P.; Tinikul, R.
    Selective determination of the catalytic cysteine pKa of two-cysteine succinic semialdehyde dehydrogenase from Acinetobacter baumannii using burst kinetics and enzyme adduct formation (2018), FEBS J., 285, 2504-2519 .
    View publication on PubMed
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Activating Compound

Activating Compound Comment Organism Structure
DTT
-
 Acinetobacter baumannii

Protein Variants

Protein Variants Comment Organism
C289A site-directed mutagenesis, inactive mutant Acinetobacter baumannii
C291A site-directed mutagenesis, the mutant shows 65% activity compared to wild-type enzyme Acinetobacter baumannii

Inhibitors

Inhibitors Comment Organism Structure
additional information no product inhibition by succinate Acinetobacter baumannii
succinate semialdehyde low substrate inhibition Acinetobacter baumannii

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
 additional information steady-state kinetic analysis. Pre-steady-state kinetics show burst kinetics for NADPH formation in SSADH, indicating that the rate-limiting step is associated with steps that occur after the hydride transfer. Detection of burst kinetics of NADPH production by pre-steady-state analysis indicates that the rate-limiting step of the AbSSADH reaction occurs after the hydride transfer step Acinetobacter baumannii
0.113
-
 NADP+ pH 8.0, 25°C Acinetobacter baumannii
0.189
-
 succinate semialdehyde pH 8.0, 25°C Acinetobacter baumannii

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
208000
-
 about, gel filtration Acinetobacter baumannii

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
succinate semialdehyde + NADP+ + H2O Acinetobacter baumannii
-
 succinate + NADPH + 2 H+
-
 ?

Organism

Organism UniProt Comment Textmining
Acinetobacter baumannii A0A0D5YDF1
-
-

Reaction

Reaction Comment Organism Reaction ID
succinate semialdehyde + NADP+ + H2O = succinate + NADPH + 2 H+ proposed reaction mechanism of AbSSADH via thiohemiacetal intermediate and thioester intermediate. Formation of Cys289 thiolate is necessary for the activity of AbSSADH. Based on two independent methods of pKa measurement, the pKa of Cys289 is 7.4-7.9. The rate-limiting step of the overall reaction of AbSSADH are the thioester intermediate hydrolysis and product liberation steps Acinetobacter baumannii
a

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
succinate semialdehyde + NADP+ + H2O
-
 Acinetobacter baumannii succinate + NADPH + 2 H+
-
 ?

Subunits

Subunits Comment Organism
homotetramer 4 * 52000, SDS-PAGE Acinetobacter baumannii

Synonyms

Synonyms Comment Organism
AbSSADH
-
 Acinetobacter baumannii
gabD
-
 Acinetobacter baumannii
SSADH
-
 Acinetobacter baumannii
succinic semialdehyde dehydrogenase
-
 Acinetobacter baumannii

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
137
-
 NADP+ pH 8.0, 25°C Acinetobacter baumannii
137
-
 succinate semialdehyde pH 8.0, 25°C Acinetobacter baumannii

pH Range

pH Minimum pH Maximum Comment Organism
additional information
-
 pH dependency of the NADP-cysteine adduct formation, pH-rate profile analysis, overview Acinetobacter baumannii

Cofactor

Cofactor Comment Organism Structure
additional information AbSSADH can use both NADP+ and NAD+ as electron acceptors but has a greater preference for NADP+. The specific activity of the enzyme with NADP+ is 10times higher than that with NAD+. Residue Ser183 is involved in cofactor binding Acinetobacter baumannii
NADP+
-
 Acinetobacter baumannii

General Information

General Information Comment Organism
evolution sequence analysis of AbSSADH reveals its high degree of sequence similarity to other enzymes in the two-cysteine GabD family Acinetobacter baumannii
metabolism the ssadh gene from Acinetobacter baumannii is part of the 4-hydroxyphenylacetate (4-HPA) degradation pathway in which SSADH converts SSA to succinic acid before entering the main metabolic TCA cycle Acinetobacter baumannii
additional information AbSSADH contains a total of five cysteine residues in one subunit (Cys175, Cys245, Cys289, Cys291 and Cys479), the enzyme has two conserved cysteines, Cys289 and Cys291. Cys289 is the active residue participating in catalysis. Method devlopment to specifically measure the active site cysteine pKa without interference from other cysteines, overview. As the magnitude of burst kinetics represents the amount of NADPH formed during the first turnover, it is directly dependent on the amount of the deprotonated form of cysteine. The pKa of Cys289 was calculated from a plot of the burst magnitude vs. pH as 7.4. The Cys289 pKa is also measured based on the ability of AbSSADH to form an NADP-cysteine adduct, which can be detected by the increase of absorbance at about 330 nm as 7.9. The lowering of the catalytic cysteine pKa by 0.6-1.0 unit renders the catalytic thiol more nucleophilic, which facilitates AbSSADH catalysis under physiological conditions. Deprotonation of the ligand or an active site residue is required for the SSADH reaction Acinetobacter baumannii

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
724.9
-
 succinate semialdehyde pH 8.0, 25°C Acinetobacter baumannii
1212.4
-
 NADP+ pH 8.0, 25°C Acinetobacter baumannii