New approach of three-dimensional crystallization of the Ca(2+)-ATPase of sarcoplasmic reticulum.
Selective extraction procedure was applied for obtaining different proteins from sarcoplasmic reticulum vesicles following the main steps of previous observations. High concentration of a nonionic detergent, such as polyoxyethylene-10-lauryl ether (C12E10) prevented the formation of Ca(2+)-ATPase crystals. It has been observed that only the 300 kDa protein could induce crystallization from among proteins being undissolved from of membrane. A modification of MacLennan's procedure--applied for ATPase precipitation from deoxycholate solubilized sarcoplasmic reticulum--has been described and an ammonium acetate precipitated Ca(2+)-ATPase was used in the experiments for increase of Ca(2+)-ATPase concentration in the crystallization process. The repeated supplementation of purified and C12E10 solubilized Ca(2+)-ATPase with ammonium acetate precipitated Ca(2+)-ATPase made possible a formation of larger and larger crystals with different periodicity.
Keresztes
T
T
Central Research Laboratory, University Medical School, Debrecen, Hungary.
Varga
S
S
Kövér
A
A
D000818:Animals; D000252:Calcium-Transporting ATPases / Q000737:chemistry* / Q000302:isolation & purification / Q000648:ultrastructure; D003460:Crystallization; D003902:Detergents; D004591:Electrophoresis, Polyacrylamide Gel; D008970:Molecular Weight; D011817:Rabbits; D012519:Sarcoplasmic Reticulum / Q000201:enzymology*; D012995:Solubility
D003902:Detergents; D000252:Calcium-Transporting ATPases