1dt1
From Proteopedia
(New page: 200px<br /><applet load="1dt1" size="450" color="white" frame="true" align="right" spinBox="true" caption="1dt1, resolution 1.8Å" /> '''THERMUS THERMOPHILUS ...) |
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| - | [[Image:1dt1.gif|left|200px]]<br /><applet load="1dt1" size=" | + | [[Image:1dt1.gif|left|200px]]<br /><applet load="1dt1" size="350" color="white" frame="true" align="right" spinBox="true" |
caption="1dt1, resolution 1.8Å" /> | caption="1dt1, resolution 1.8Å" /> | ||
'''THERMUS THERMOPHILUS CYTOCHROME C552 SYNTHESIZED BY ESCHERICHIA COLI'''<br /> | '''THERMUS THERMOPHILUS CYTOCHROME C552 SYNTHESIZED BY ESCHERICHIA COLI'''<br /> | ||
==Overview== | ==Overview== | ||
| - | We describe the design of Escherichia coli cells that synthesize a | + | We describe the design of Escherichia coli cells that synthesize a structurally perfect, recombinant cytochrome c from the Thermus thermophilus cytochrome c552 gene. Key features are (1) construction of a plasmid-borne, chimeric cycA gene encoding an Escherichia coli-compatible, N-terminal signal sequence (MetLysIleSerIleTyrAlaThrLeu AlaAlaLeuSerLeuAlaLeuProAlaGlyAla) followed by the amino acid sequence of mature Thermus cytochrome c552; and (2) coexpression of the chimeric cycA gene with plasmid-borne, host-specific cytochrome c maturation genes (ccmABCDEFGH). Approximately 1 mg of purified protein is obtained from 1 L of culture medium. The recombinant protein, cytochrome rsC552, and native cytochrome c552 have identical redox potentials and are equally active as electron transfer substrates toward cytochrome ba3, a Thermus heme-copper oxidase. Native and recombinant cytochromes c were compared and found to be identical using circular dichroism, optical absorption, resonance Raman, and 500 MHz 1H-NMR spectroscopies. The 1.7 A resolution X-ray crystallographic structure of the recombinant protein was determined and is indistinguishable from that reported for the native protein (Than, ME, Hof P, Huber R, Bourenkov GP, Bartunik HD, Buse G, Soulimane T, 1997, J Mol Biol 271:629-644). This approach may be generally useful for expression of alien cytochrome c genes in E. coli. |
==About this Structure== | ==About this Structure== | ||
| - | 1DT1 is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Thermus_thermophilus Thermus thermophilus] with HEM as [http://en.wikipedia.org/wiki/ligand ligand]. Full crystallographic information is available from [http:// | + | 1DT1 is a [http://en.wikipedia.org/wiki/Single_protein Single protein] structure of sequence from [http://en.wikipedia.org/wiki/Thermus_thermophilus Thermus thermophilus] with <scene name='pdbligand=HEM:'>HEM</scene> as [http://en.wikipedia.org/wiki/ligand ligand]. Full crystallographic information is available from [http://oca.weizmann.ac.il/oca-bin/ocashort?id=1DT1 OCA]. |
==Reference== | ==Reference== | ||
| Line 15: | Line 15: | ||
[[Category: Ai, J.]] | [[Category: Ai, J.]] | ||
[[Category: Chen, Y.]] | [[Category: Chen, Y.]] | ||
| - | [[Category: Fee, J | + | [[Category: Fee, J A.]] |
[[Category: Gomez-Moran, E.]] | [[Category: Gomez-Moran, E.]] | ||
| - | [[Category: Hill, M | + | [[Category: Hill, M J.]] |
[[Category: Loehr, T.]] | [[Category: Loehr, T.]] | ||
| - | [[Category: McRee, D | + | [[Category: McRee, D E.]] |
[[Category: Sridhar, V.]] | [[Category: Sridhar, V.]] | ||
[[Category: Stura, E.]] | [[Category: Stura, E.]] | ||
[[Category: Thony-Meyer, L.]] | [[Category: Thony-Meyer, L.]] | ||
| - | [[Category: Williams, P | + | [[Category: Williams, P A.]] |
[[Category: HEM]] | [[Category: HEM]] | ||
[[Category: cytochrome c552]] | [[Category: cytochrome c552]] | ||
[[Category: thermus thermophilus]] | [[Category: thermus thermophilus]] | ||
| - | ''Page seeded by [http:// | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Feb 21 12:20:08 2008'' |
Revision as of 10:20, 21 February 2008
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THERMUS THERMOPHILUS CYTOCHROME C552 SYNTHESIZED BY ESCHERICHIA COLI
Overview
We describe the design of Escherichia coli cells that synthesize a structurally perfect, recombinant cytochrome c from the Thermus thermophilus cytochrome c552 gene. Key features are (1) construction of a plasmid-borne, chimeric cycA gene encoding an Escherichia coli-compatible, N-terminal signal sequence (MetLysIleSerIleTyrAlaThrLeu AlaAlaLeuSerLeuAlaLeuProAlaGlyAla) followed by the amino acid sequence of mature Thermus cytochrome c552; and (2) coexpression of the chimeric cycA gene with plasmid-borne, host-specific cytochrome c maturation genes (ccmABCDEFGH). Approximately 1 mg of purified protein is obtained from 1 L of culture medium. The recombinant protein, cytochrome rsC552, and native cytochrome c552 have identical redox potentials and are equally active as electron transfer substrates toward cytochrome ba3, a Thermus heme-copper oxidase. Native and recombinant cytochromes c were compared and found to be identical using circular dichroism, optical absorption, resonance Raman, and 500 MHz 1H-NMR spectroscopies. The 1.7 A resolution X-ray crystallographic structure of the recombinant protein was determined and is indistinguishable from that reported for the native protein (Than, ME, Hof P, Huber R, Bourenkov GP, Bartunik HD, Buse G, Soulimane T, 1997, J Mol Biol 271:629-644). This approach may be generally useful for expression of alien cytochrome c genes in E. coli.
About this Structure
1DT1 is a Single protein structure of sequence from Thermus thermophilus with as ligand. Full crystallographic information is available from OCA.
Reference
Integrity of thermus thermophilus cytochrome c552 synthesized by Escherichia coli cells expressing the host-specific cytochrome c maturation genes, ccmABCDEFGH: biochemical, spectral, and structural characterization of the recombinant protein., Fee JA, Chen Y, Todaro TR, Bren KL, Patel KM, Hill MG, Gomez-Moran E, Loehr TM, Ai J, Thony-Meyer L, Williams PA, Stura E, Sridhar V, McRee DE, Protein Sci. 2000 Nov;9(11):2074-84. PMID:11152119
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