X-ray structure of putative acyl-ACP desaturase DesA2 from Mycobacterium tuberculosis H37Rv

Dyer, David H; Lyle, Karen S; Rayment, Ivan; Fox, Brian G

Title: X-ray structure of putative acyl-ACP desaturase DesA2 from Mycobacterium tuberculosis H37Rv

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Abstract

Genome sequencing showed that two proteins in Mycobacterium tuberculosis H37Rv contain the metal binding motif (D/E)X{sub 2}HX{sub {approx}100}(D/E)X{sub 2}H characteristic of the soluble diiron enzyme superfamily. These putative acyl-ACP desaturase genes desA1 and desA2 were cloned from genomic DNA and expressed in Escherichia coli BL21(DE3). DesA1 was found to be insoluble, but in contrast, DesA2 was a soluble protein amenable to biophysical characterization. Here, we report the 2.0 {angstrom} resolution X-ray structure of DesA2 determined by multiple anomalous dispersion (MAD) phasing from a Se-met derivative and refinement against diffraction data obtained on the native protein. The X-ray structure shows that DesA2 is a homodimeric protein with a four-helix bundle core flanked by five additional helices that overlay with 192 structurally equivalent amino acids in the structure of stearoyl-ACP {Delta}9 desaturase from castor plant with an rms difference 1.42 {angstrom}. In the DesA2 crystals, one metal (likely Mn from the crystallization buffer) was bound in high occupancy at the B-site of the conserved metal binding motif, while the A-site was not occupied by a metal ion. Instead, the amino group of Lys-76 occupied this position. The relationships between DesA2 and known diiron enzymes are discussed.

Authors:

Dyer, David H; Lyle, Karen S; Rayment, Ivan; Fox, Brian G ^[1]

Publication Date:: Tue Jul 13 00:00:00 EDT 2010

Research Org.:: Argonne National Lab. (ANL), Argonne, IL (United States). Advanced Photon Source (APS)

Sponsoring Org.:: USDOE

OSTI Identifier:: 1008567

Resource Type:: Journal Article

Journal Name:: Protein Sci.

Additional Journal Information:: Journal Volume: 14; Journal Issue: (6) ; 2005

Country of Publication:: United States

Language:: ENGLISH

Subject:: 59 BASIC BIOLOGICAL SCIENCES; 60 APPLIED LIFE SCIENCES; AMINO ACIDS; CASTOR; CRYSTALLIZATION; DIFFRACTION; DNA; ENZYMES; ESCHERICHIA COLI; GENES; MYCOBACTERIUM TUBERCULOSIS; PROTEINS; RESOLUTION

Citation Formats


                    Dyer, David H, Lyle, Karen S, Rayment, Ivan, and Fox, Brian G. X-ray structure of putative acyl-ACP desaturase DesA2 from Mycobacterium tuberculosis H37Rv.  United States: N. p., 2010. 
        Web.

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                    Dyer, David H, Lyle, Karen S, Rayment, Ivan, & Fox, Brian G. X-ray structure of putative acyl-ACP desaturase DesA2 from Mycobacterium tuberculosis H37Rv.  United States.

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                    Dyer, David H, Lyle, Karen S, Rayment, Ivan, and Fox, Brian G. 2010.  
        "X-ray structure of putative acyl-ACP desaturase DesA2 from Mycobacterium tuberculosis H37Rv".  United States.

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@article{osti_1008567,

  title        = {X-ray structure of putative acyl-ACP desaturase DesA2 from Mycobacterium tuberculosis H37Rv},

  author       = {Dyer, David H and Lyle, Karen S and Rayment, Ivan and Fox, Brian G},

  abstractNote = {Genome sequencing showed that two proteins in Mycobacterium tuberculosis H37Rv contain the metal binding motif (D/E)X{sub 2}HX{sub {approx}100}(D/E)X{sub 2}H characteristic of the soluble diiron enzyme superfamily. These putative acyl-ACP desaturase genes desA1 and desA2 were cloned from genomic DNA and expressed in Escherichia coli BL21(DE3). DesA1 was found to be insoluble, but in contrast, DesA2 was a soluble protein amenable to biophysical characterization. Here, we report the 2.0 {angstrom} resolution X-ray structure of DesA2 determined by multiple anomalous dispersion (MAD) phasing from a Se-met derivative and refinement against diffraction data obtained on the native protein. The X-ray structure shows that DesA2 is a homodimeric protein with a four-helix bundle core flanked by five additional helices that overlay with 192 structurally equivalent amino acids in the structure of stearoyl-ACP {Delta}9 desaturase from castor plant with an rms difference 1.42 {angstrom}. In the DesA2 crystals, one metal (likely Mn from the crystallization buffer) was bound in high occupancy at the B-site of the conserved metal binding motif, while the A-site was not occupied by a metal ion. Instead, the amino group of Lys-76 occupied this position. The relationships between DesA2 and known diiron enzymes are discussed.},

  doi          = {},

  url          = {https://www.osti.gov/biblio/1008567},
  journal      = {Protein Sci.},
number       = (6) ; 2005,

  volume       = 14,

  place        = {United States},

  year         = {Tue Jul 13 00:00:00 EDT 2010},

  month        = {Tue Jul 13 00:00:00 EDT 2010}

}

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