Activating Compound | Comment | Organism | Structure |
---|---|---|---|
additional information | the catalytic efficiency of MICAL3 increases on adding F-actin only when the CH domain is available | Homo sapiens |
Cloned (Comment) | Organism |
---|---|
gene MICAL3, hMICAL3FMOCH, hMICAL3FMO, and hMICAL3FMODCHELTA213,530 are cloned into the pET-28b plasmid, recombinant overexpression of His-tagged wild-type and mutant enzymes in Escherichia coli strain Rosetta 2 pLysS | Homo sapiens |
Crystallization (Comment) | Organism |
---|---|
MICAL3 has an FAD/NADP-binding Rossmann-fold domain for monooxygenase activity. The flavin-containing monooxygenase (FMO) and calponin-homology (CH) domains of both MICAL3 and MICAL1 are highly similar in structure, but a different relative position of the calponin-homology domain in the asymmetric unit | Homo sapiens |
purified recombinant His-tagged wild-type and mutant MICAL3 variants, sitting drop vapor diffusion method, method optimization, mixing of 500 nl of 25 mg/ml protein in 50 mM Tris, pH 8.5, 100 mM NaCl, 1 mM 1,4-dithiothreitol, 1% glycerol, with 500 nl of crystallization solution containing 0.1 M bicine-NaOH, pH 9.2, 7% v/v MPD, one day, X-ray diffraction structure determination and analysis at 1.9-2.3 A resolution | Homo sapiens |
Protein Variants | Comment | Organism |
---|---|---|
E213G | site-directed mutagenesis in the FMO domain | Homo sapiens |
E213G/R530G | site-directed mutagenesis | Homo sapiens |
additional information | construction of enzyme mutants hMCAL3FMOCH, hMICAL3FMO and hMICAL3FMOCHDELTA213,530. The truncated form containing the FMO and CH domains is much more soluble than the full-length form but still retains catalytic activity for F-actin disassembly | Homo sapiens |
R530G | site-directed mutagenesis in the CH domain | Homo sapiens |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
additional information | - |
additional information | Michaelis-Menten kinetics | Homo sapiens | |
0.2669 | - |
[F-actin]-L-methionine | pH 8.0, 25°C, recombinant hMICAL3FMOCH | Homo sapiens | |
0.5438 | - |
[F-actin]-L-methionine | recombinant MICAL3 containing the FMO and CH domains, pH 7.5, 25°C | Homo sapiens | |
0.8172 | - |
[F-actin]-L-methionine | recombinant MICAL3 lacking the CH domain, pH 7.5, 25°C | Homo sapiens | |
1.264 | - |
[F-actin]-L-methionine | pH 8.0, 25°C, recombinant hMICAL3FMO | Homo sapiens | |
1.438 | - |
[F-actin]-L-methionine | pH 8.0, 25°C, recombinant hMICAL3FMOCHDELTA213,530 | Homo sapiens |
Localization | Comment | Organism | GeneOntology No. | Textmining |
---|---|---|---|---|
nucleus | - |
Homo sapiens | 5634 | - |
Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
---|---|---|---|
79400 | - |
recombinant His-tagged enzyme, gel filtration | Homo sapiens |
Natural Substrates | Organism | Comment (Nat. Sub.) | Natural Products | Comment (Nat. Pro.) | Rev. | Reac. |
---|---|---|---|---|---|---|
[F-actin]-L-methionine + NADPH + O2 + H+ | Homo sapiens | - |
[F-actin]-L-methionine-(R)-S-oxide + NADP+ + H2O | - |
? |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Homo sapiens | Q7RTP6 | - |
- |
Homo sapiens | Q7RTP6 | isoform MICAL3 | - |
Purification (Comment) | Organism |
---|---|
recombinant His-tagged wild-type and mutant enzymes from Escherichia coli strain Rosetta 2 pLysS by nickel affinity chromatography, ultrafiltration, and gel filtration | Homo sapiens |
Source Tissue | Comment | Organism | Textmining |
---|---|---|---|
skeletal muscle | - |
Homo sapiens | - |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
additional information | MICAL has additionally NADPH oxidase activity that underlies F-actin disassembly simultaneously with the oxidation of NADPH. For MICAL3 containing the FMO and CH domains, the kcat/Km ratio for NADPH oxidase activity increases dramatically on adding F-actin, while the kcat/Km value for MICAL3 lacking the CH domain changes little | Homo sapiens | ? | - |
- |
|
[F-actin]-L-methionine + NADPH + O2 + H+ | - |
Homo sapiens | [F-actin]-L-methionine-(R)-S-oxide + NADP+ + H2O | - |
? | |
[F-actin]-L-methionine + NADPH + O2 + H+ | usage of F-actin from rabbit skeletal muscle purified from G-actin by ultracentrifugation | Homo sapiens | [F-actin]-L-methionine-(R)-S-oxide + NADP+ + H2O | - |
? |
Subunits | Comment | Organism |
---|---|---|
homodimer | 2 * 40000, about, SDS-PAGE | Homo sapiens |
More | human MICAL3 contains the flavin-containing monooxygenase (FMO) and calponin-homology (CH) domains, and has an FAD/NADP-binding Rossmann-fold domain for monooxygenase activity like MICAL1, structure comparisons of isozymes MICAL3 and MICAL1. The FMO and CH domains of both isozymes MICAL3 and MICAL1 are highly similar in structure, but superimposition of the two structures shows a different relative position of the CH domain in the asymmetric unit. The catalytic efficiency of MICAL3 dramatically increases on adding F-actin only when the CH domain is available | Homo sapiens |
Synonyms | Comment | Organism |
---|---|---|
MICAL3 | - |
Homo sapiens |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
25 | - |
assay at | Homo sapiens |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.0006 | - |
[F-actin]-L-methionine | pH 8.0, 25°C, recombinant hMICAL3FMOCH | Homo sapiens | |
0.0024 | - |
[F-actin]-L-methionine | recombinant MICAL3 lacking the CH domain, pH 7.5, 25°C | Homo sapiens | |
0.0031 | - |
[F-actin]-L-methionine | pH 8.0, 25°C, recombinant hMICAL3FMO | Homo sapiens | |
0.0033 | - |
[F-actin]-L-methionine | pH 8.0, 25°C, recombinant hMICAL3FMOCHDELTA213,530 | Homo sapiens | |
0.0169 | - |
[F-actin]-L-methionine | recombinant MICAL3 containing the FMO and CH domains, pH 7.5, 25°C | Homo sapiens |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
8 | - |
assay at | Homo sapiens |
Cofactor | Comment | Organism | Structure |
---|---|---|---|
FAD | - |
Homo sapiens | |
NADPH | - |
Homo sapiens |
General Information | Comment | Organism |
---|---|---|
malfunction | the catalytic efficiency of MICAL3 increases on adding F-actin only when the CH domain is available. But this does not occur when two residues, Glu213 and Arg530, are mutated in the FMO and CH domains, respectively | Homo sapiens |
additional information | the catalytic efficiency of MICAL3 increases on adding F-actin only when the CH domain is available. MICAL3 is structurally highly similar to isozyme MICAL1, which suggests that they may adopt the same catalytic mechanism, but the difference in the relative position of the CH domain produces a difference in F-actin substrate specificity. Interaction analysis of the binding site between the CH domain and the FMO domain in human MICAL3, modeling, overview. The FMO-CH interaction in hMICAL3 is required to increase the catalytic efficiency by conferring specific binding to F-actin. The FMO domain that exhibits monooxygenase activity is localized at the N-terminus of MICAL and is highly conserved among species. The CH domain that is usually found in actin binding proteins is adjacent to the FMO domain and is also highly conserved. CH domains are classified into three types: types 1, 2, and 3. Whereas type 3 CH domains are mainly found in regulatory proteins associated with muscle contraction and signaling proteins, type 1 and 2 CH domains are usually found in cytoskeletal proteins. MICALs have a typical type 2 CH domain | Homo sapiens |
physiological function | MICAL isozymes are involved in actin cytoskeleton reorganization through methionine oxidation. The enzyme functions in F-actin disassembly | Homo sapiens |
kcat/KM Value [1/mMs-1] | kcat/KM Value Maximum [1/mMs-1] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.00225 | - |
[F-actin]-L-methionine | pH 8.0, 25°C, recombinant hMICAL3FMOCH | Homo sapiens | |
0.0023 | - |
[F-actin]-L-methionine | pH 8.0, 25°C, recombinant hMICAL3FMOCHDELTA213,530 | Homo sapiens | |
0.00245 | - |
[F-actin]-L-methionine | pH 8.0, 25°C, recombinant hMICAL3FMO | Homo sapiens | |
0.0029 | - |
[F-actin]-L-methionine | recombinant MICAL3 lacking the CH domain, pH 7.5, 25°C | Homo sapiens | |
0.031 | - |
[F-actin]-L-methionine | recombinant MICAL3 containing the FMO and CH domains, pH 7.5, 25°C | Homo sapiens |