Characterization of Zn(II)-responsive ribosomal proteins YkgM and L31 in E. coli
- 作者：M. Patrick Hensley ; Thusitha S. Gunasekera ; J. Allen Easton ; Tara K. Sigdel ; Stacy A. Sugarbaker ; Lindsey Klingbeil ; Robert M. Breece ; David L. Tierney ; Michael W. Crowder ; crowdemw@muohio.edu
- 关键词：Ribosomal protein L31 ; EXAFS ; Zn(II) homeostasis ; DNA microarrays
- 刊名：Journal of Inorganic Biochemistry
- 出版年：2012
- 期刊代码：53_01620134
- 类别：ch
- 出版时间：June, 2012
- 卷：111
- 期：Complete
- 页码：164-172
- 文件大小：581 K
摘要
RT-PCR and DNA microarrays were used to probe for Zn(II)-responsive genes in E. coli cells that were made Zn(II) deficient. Microarray data revealed 114 genes were significantly up-regulated and 146 genes were significantly down-regulated in Zn(II) deficient conditions. The three most up-regulated genes were (1) znuA, which encodes for a periplasmic protein known to be involved with Zn(II) import, (2) yodA, which encodes for a periplasmic protein with unknown function, and (3) ykgM, which encodes for a ribosomal protein that is thought to be a paralog of ribosomal protein L31. YodA was over-expressed and purified as a maltose binding protein (MBP) fusion protein and shown to tightly bind 4 equivalents of Zn(II). Metal analyses showed that MBP-YkgM does not bind Zn(II). On the other hand, MBP-L31 tightly binds 1 equivalent of Zn(II). EXAFS studies on MBP-L31 suggest a ligand field of 1 histidine, 1 cysteine, and 2 additional N/O scatterers. Site-directed mutagenesis studies suggest that Cys16 coordinates Zn(II) in MBP-L31 and that the other three cysteines do not bind metal. These results are discussed in light of Zn(II) starvation model that has been postulated for B. subtilis.