title: Putative copper- and zinc-binding motifs in Streptococcus pneumoniae identified by immobilized metal affinity chromatography and mass spectrometry.
abstract: The aim of metalloproteomics is to identify and characterize putative metal-binding proteins and metal-binding motifs. In this study, we performed a systematical metalloproteomic analysis on Streptococcus pneumoniae through the combined use of efficient immobilized metal affinity chromatography enrichment and high-accuracy linear ion trap-Orbitrap MS to identify metal-binding proteins and metal-binding peptides. In total, 232 and 166 putative metal-binding proteins were respectively isolated by Cu- and Zn-immobilized metal affinity chromatography columns, in which 133 proteins were present in both preparations. The putative metalloproteins are mainly involved in protein, nucleotide and carbon metabolisms, oxidation and cell cycle regulation. Based on the sequence of the putative Cu- and Zn-binding peptides, putative Cu-binding motifs were identified: H(X)mH (m=0-11), C(X)(2) C, C(X)nH (n=2-4, 6, 9), H(X)iM (i=0-10) and M(X)tM (t=8 or 12), while putative Zn-binding motifs were identified as follows: H(X)mH (m=1-12), H(X)iM (i=0-12), M(X)tM (t=0, 3 and 4), C(X)nH (n=1, 2, 7, 10 and 11). Equilibrium dialysis and inductively coupled plasma-MS experiments confirmed that the artificially synthesized peptides harboring differential identified metal-binding motifs interacted directly with the metal ions. The metalloproteomic study presented here suggests that the comparably large size and diverse functions of the S. pneumoniae metalloproteome may play important roles in various biological processes and thus contribute to the bacterial pathologies.
authors.affiliation: Institute of Life and Health Engineering/National Engineering and Research Center of Genetic Medicine, Jinan University, Guangzhou, P R China. tsunxs@jnu.edu.cn
authors.forename: Xuesong
authors.initials: X
authors.lastname: Sun
authors.forename: Chuan-le
authors.initials: CL
authors.lastname: Xiao
authors.forename: Ruiguang
authors.initials: R
authors.lastname: Ge
authors.forename: Xingfeng
authors.initials: X
authors.lastname: Yin
authors.forename: Hui
authors.initials: H
authors.lastname: Li
authors.forename: Nan
authors.initials: N
authors.lastname: Li
authors.forename: Xiaoyan
authors.initials: X
authors.lastname: Yang
authors.forename: Ying
authors.initials: Y
authors.lastname: Zhu
authors.forename: Xiang
authors.initials: X
authors.lastname: He
authors.forename: Qing-Yu
authors.initials: QY
authors.lastname: He
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000032:analysis / Q000737:chemistry* / Q000378:metabolism; D002352:Carrier Proteins / Q000032:analysis / Q000737:chemistry / Q000378:metabolism; D002846:Chromatography, Affinity; D003300:Copper / Q000737:chemistry / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D042541:Intracellular Space / Q000737:chemistry / Q000378:metabolism; D013058:Mass Spectrometry; D008667:Metalloproteins / Q000032:analysis / Q000737:chemistry* / Q000378:metabolism; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000032:analysis / Q000737:chemistry / Q000378:metabolism; D020543:Proteome / Q000032:analysis / Q000737:chemistry / Q000378:metabolism; D040901:Proteomics; D013296:Streptococcus pneumoniae / Q000737:chemistry / Q000378:metabolism*; D015032:Zinc / Q000737:chemistry / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D002352:Carrier Proteins; D008667:Metalloproteins; D010446:Peptide Fragments; D020543:Proteome; D003300:Copper; D015032:Zinc