Purification and some properties of a blue copper protein from Methylobacillus sp. strain SK1 DSM 8269

Si Wook Kim, Jeong Ae Kim, Eungbin Kim, Young Tae Ro, Taeksun Song, Young Min Kim

Research output: Contribution to journalArticle

Abstract

A blue protein was purified from the Methylobacillus sp. strain SK1 that is grown on methanol in the presence of copper ion. This protein was found to be a monomer with a molecular weight of 13,500. The Isoelectric point of the protein was estimated to be 8.8. The spectrum of the protein that was treated with ferricyanide showed a broad peak around 620 nm, but that of the dithionite-treated protein revealed no peaks. It contained 0.83 mol of EDTA-stable copper per mol protein. Under air, the protein accelerated the inactivation of methanol dehydrogenase (MDH). The protein was reducible by phenazine methosulfate or by active MDH that was prepared from cells that were grown in the absence of added copper, but not by methanol, dichlorophenol indophenol, or inactive MDH that was prepared from cells that were grown in the presence of added copper. It was also reducible by active MDH in the presence of methanol. The absorption peak at 340 nm of the active MDH disappeared after the enzyme was treated with ferricyanide, hydrogen peroxide, or the purified blue protein. The inactive MDH also showed no peak at 340 nm. The 340-nm peak was not recovered after incubation of the inactive MDH and blue protein-treated active MDH with dithionite or methanol. The inactive MDH and blue protein-treated active MDH co-migrated with the active MDH preparation on nondenaturing polyacrylamide gel, and contained two non-identical subunits with molecular weights that were identical to those of the active MDH. The N-terminal amino acid sequence of the protein was Ala-Gly-Cys-Ser-Val-Asp-Val-Glu-Ala-Asn-Asp- Ala-Met-Gln-Phe. An analysis of the amino acid composition revealed that the protein contained no tryptophan. It contained three cysteines per mol protein. The blue protein in Methylobacillus sp. strain SK1 was produced only in the cells that were grown in the copper-supplemented medium.

Original languageEnglish
Pages (from-to)214-223
Number of pages10
JournalMolecules and cells
Volume14
Issue number2
Publication statusPublished - 2002 Oct 1

All Science Journal Classification (ASJC) codes

  • Molecular Biology
  • Cell Biology

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