Reaktionsgleichung
P([O-])([O-])([O-])=O.[CH3:6][C@@H:7]1[O:11][C@@H:10]([O:12][C@H:13]2[C@H:18]([OH:19])[C@@H:17]([OH:20])[C@H:16]([N:21]=[C:22]([NH2:24])[NH2:23])[C@@H:15]([OH:25])[C@@H:14]2[N:26]=[C:27]([NH2:29])[NH2:28])[C@H:9]([O:30][C@@H:31]2[O:36][C@@H:35]([CH2:37][OH:38])[C@H:34]([OH:39])[C@@H:33]([OH:40])[C@@H:32]2[NH:41][CH3:42])[C@@:8]1([OH:45])[CH:43]=[O:44].OS(O)(=O)=O>>[CH3:6][C@@H:7]1[O:11][C@@H:10]([O:12][C@H:13]2[C@H:18]([OH:19])[C@@H:17]([OH:20])[C@H:16]([NH:21][C:22]([NH2:24])=[NH:23])[C@@H:15]([OH:25])[C@@H:14]2[NH:26][C:27]([NH2:29])=[NH:28])[C@H:9]([O:30][C@@H:31]2[O:36][C@@H:35]([CH2:37][OH:38])[C@H:34]([OH:39])[C@@H:33]([OH:40])[C@@H:32]2[NH:41][CH3:42])[C@@:8]1([OH:45])[CH:43]=[O:44]Komponenten
Versuchsvorschrift
The thus-obtained cell was suspended in 600 ml of a 0.1M phosphate buffer solution, and broken by the use of a Dyno-mill. After the removal of insoluble materials by centrifugal separation, a crude extract containing adenylate kinase was obtained. Then, 200 ml of a 10% streptomycin sulfate solution was added to 400 ml of the crude extract. Precipitates formed were removed by centrifugal separation to thereby obtain a streptomycin supernatant liquid. This supernatant liquid was subjected to ammonium sulfate fractionation, and a fraction from 30% saturation (4° C.) to 60% saturation (4° C.) was obtained. This fraction was dissolved in a 50 mM Tris-hydrochloric acid buffer solution (pH 8.0) and passed through a DEAE-Sephadex column which had been equilibrated with the same buffer solution as above, and thereafter it was eluted with a solution prepared by adding sodium chloride to the above buffer solution. Near a concentration of sodium chloride of 0.2M, the desired adenylate kinase was eluted. The thus-eluted fraction was subjected to hydroxyapatite column chromatography under the same conditions as in Example 1, passed through a Sephadex G-75 column, and eluted with a 30 mM Tris-hydrochloric acid buffer solution (pH 8.0) containing 0.1M sodium chloride to thereby obtain an adenylate kinase sample which provided a single band by acrylamide disc electrophoresis as in the case of Example 1. Furthermore, as in the case of Example 1, the adenylate kinase sample provided a single peak at a molecular weight of about 22,000 as determined by Sephadex G-100 chromatography.
Aufarbeitung
- customAfter the removal of insoluble materials
- customby centrifugal separation
- extractiona crude extract
- additioncontaining adenylate kinase
- customwas obtained
- extractionextract
- customPrecipitates
- customformed
- customwere removed by centrifugal separation
Quellen und Nachweise
- ORD-Eintrag
- ord-9e96ee3ffb2b4b19974f6f92d8b2dd7c
- Quelldatensatz
- uspto-grants-1986_04
- Patentnummer
- US04584272
Verknüpfte Notizen
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