Ni-chelate-affinity purification and crystallization of the yeast mitochondrial F1-ATPase

被引：20

作者：

Mueller, DM ^{[1
]}

Puri, N

Kabaleeswaran, V

Terry, C

Leslie, AGW

Walker, JE

机构：

[1] Chicago Med Sch, Dept Biochem & Mol Biol, Chicago, IL USA

[2] MRC, Dunn Human Nutr Unit, Cambridge, England

[3] MRC, Mol Biol Lab, Cambridge CB2 2QH, England

来源：

PROTEIN EXPRESSION AND PURIFICATION | 2004年 / 37卷 / 02期

关键词：

mitochondrion; ATPase; ATP synthase; crystallization;

D O I：

10.1016/j.pep.2004.06.035

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

The yeast mitochondrial ATPase has been genetically modified to include a HiS(6) Ni-affinity tag on the amino end of the mature P-subunit. The modified beta-subunit is imported into the mitochondrion, properly processed to the mature form, and assembled into a mature and fully active ATP synthase. The F-1-ATPase has been purified from submitochondrial particles after release from the membrane with chloroform, followed by Ni-chelate-affinity and gel filtration chromatography. The final enzyme is a homogeneous preparation with full activity and no apparent degradation products. This enzyme preparation has been used to obtain crystals that diffract to better than 2.8 Angstrom resolution. (C) 2004 Elsevier Inc. All rights reserved.

引用

页码：479 / 485

页数：7

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