| http://purl.uniprot.org/citations/15358367 | http://www.w3.org/1999/02/22-rdf-syntax-ns#type | http://purl.uniprot.org/core/Journal_Citation |
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| http://purl.uniprot.org/citations/15358367 | http://www.w3.org/2000/01/rdf-schema#comment | "Dimethylglycine dehydrogenase (Me(2)GlyDH) is a mitochondrial enzyme that catalyzes the oxidative demethylation of dimethylglycine to sarcosine. The enzyme requires flavin adenine dinucleotide (FAD), which is covalently bound to the apoprotein via a histidyl(N3)-(8alpha)FAD linkage. In the present study, the mature form of rat Me(2)GlyDH has been over-expressed in Escherichia coli as an N-terminally 6-His-tagged fusion protein. The over-expressed protein distributed almost equally between the soluble and insoluble (inclusion bodies) cell fraction. By applying the soluble cell lysate to a nickel-chelating column, two fractions were eluted, both containing a nearly homogeneous protein with a molecular mass of 93 kDa, on SDS-PAGE. The first protein fraction was identified by Western blotting analysis as the covalently flavinylated Me(2)GlyDH. It showed optical properties and specific activity (240 nmol/min/mg protein) similar to those of the native holoenzyme. The second fraction was identified as an underflavinylated (apo-) form of Me(2)GlyDH, with a 70% lower specific activity. The recombinant holoenzyme exhibited optimal activity at pH 8.5, an activation energy of about 80 kJ/mol, and two KM values for N,N-dimethylglycine (KM1 = 0.05 mM and KM2 = 9.4 mM), as described for the native holoenzyme. Starting from the inclusion bodies, the unfolded flavinylated enzyme was solubilized by SDS treatment and refolded by an 80-fold dilution step, with a reactivation yield of 50-60%."xsd:string |
| http://purl.uniprot.org/citations/15358367 | http://purl.org/dc/terms/identifier | "doi:10.1016/j.pep.2004.06.011"xsd:string |
| http://purl.uniprot.org/citations/15358367 | http://purl.uniprot.org/core/author | "Indiveri C."xsd:string |
| http://purl.uniprot.org/citations/15358367 | http://purl.uniprot.org/core/author | "Pochini L."xsd:string |
| http://purl.uniprot.org/citations/15358367 | http://purl.uniprot.org/core/author | "Brandsch R."xsd:string |
| http://purl.uniprot.org/citations/15358367 | http://purl.uniprot.org/core/author | "Barile M."xsd:string |
| http://purl.uniprot.org/citations/15358367 | http://purl.uniprot.org/core/author | "Brizio C."xsd:string |
| http://purl.uniprot.org/citations/15358367 | http://purl.uniprot.org/core/author | "Bufano D."xsd:string |
| http://purl.uniprot.org/citations/15358367 | http://purl.uniprot.org/core/date | "2004"xsd:gYear |
| http://purl.uniprot.org/citations/15358367 | http://purl.uniprot.org/core/name | "Protein Expr Purif"xsd:string |
| http://purl.uniprot.org/citations/15358367 | http://purl.uniprot.org/core/pages | "434-442"xsd:string |
| http://purl.uniprot.org/citations/15358367 | http://purl.uniprot.org/core/title | "Over-expression in Escherichia coli, functional characterization and refolding of rat dimethylglycine dehydrogenase."xsd:string |
| http://purl.uniprot.org/citations/15358367 | http://purl.uniprot.org/core/volume | "37"xsd:string |
| http://purl.uniprot.org/citations/15358367 | http://www.w3.org/2004/02/skos/core#exactMatch | http://purl.uniprot.org/pubmed/15358367 |
| http://purl.uniprot.org/citations/15358367 | http://www.w3.org/2004/02/skos/core#exactMatch | http://purl.uniprot.org/pubmed/15358367 |
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