A novel approach for identifying the heme-binding proteins from mouse tissues.
Heme is a key cofactor in aerobic life, both in eukaryotes and prokaryotes. Because of the high reactivity of ferrous protoporphyrin IX, the reactions of heme in cells are often carried out through heme-protein complexes. Traditionally studies of heme-binding proteins have been approached on a case by case basis, thus there is a limited global view of the distribution of heme-binding proteins in different cells or tissues. The procedure described here is aimed at profiling heme-binding proteins in mouse tissues sequentially by 1) purification of heme-binding proteins by heme-agarose, an affinity chromatographic resin; 2) isolation of heme-binding proteins by SDS-PAGE or two-dimensional electrophoresis; 3) identification of heme-binding proteins by mass spectrometry. In five mouse tissues, over 600 protein spots were visualized on 2-DE gel stained by Commassie blue and 154 proteins were identified by MALDI-TOF, in which most proteins belong to heme related. This methodology makes it possible to globally characterize the heme-binding proteins in a biological system.
Li
Xiaolei
X
Beijing Genomics Institute, Chinese Academy of Sciences, Beijing 101300, China.
Wang
Xiaoshan
X
Zhao
Kang
K
Zhou
Zhengfeng
Z
Zhao
Caifeng
C
Yan
Ren
R
Lin
Liang
L
Lei
Tingting
T
Yin
Jianning
J
Wang
Rong
R
Sun
Zhongsheng
Z
Xu
Zuyuan
Z
Bao
Jingyue
J
Zhang
Xiuqing
X
Feng
Xiaoli
X
Liu
Siqi
S
D000818:Animals; D002352:Carrier Proteins / Q000096:biosynthesis* / Q000235:genetics*; D015180:Electrophoresis, Gel, Two-Dimensional; D004591:Electrophoresis, Polyacrylamide Gel; D006418:Heme / Q000737:chemistry; D000081062:Heme-Binding Proteins; D006420:Hemeproteins / Q000096:biosynthesis* / Q000235:genetics*; D013058:Mass Spectrometry; D051379:Mice; D008813:Mice, Inbred ICR; D011485:Protein Binding; D011506:Proteins / Q000737:chemistry; D020543:Proteome; D040901:Proteomics / Q000379:methods; D012685:Sepharose / Q000737:chemistry; D014018:Tissue Distribution
D002352:Carrier Proteins; D000081062:Heme-Binding Proteins; D006420:Hemeproteins; D011506:Proteins; D020543:Proteome; D006418:Heme; D012685:Sepharose