Sandbox Reserved 770: Difference between revisions
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'''A) Expression and Purification'''. Escherichia coli cells containing plasmid pEAL (PAL inserted into pET-24a) were grown in media methionine | '''A) Expression and Purification'''. Escherichia coli cells containing plasmid pEAL (PAL inserted into pET-24a) were grown in media methionine | ||
'''B) Protein Isolation by Centrifugation''' | '''B) Protein Isolation by Centrifugation''' | ||
'''C) Protein was purified by three chromatography steps''': | '''C) Protein was purified by three chromatography steps''': | ||
(1) Anion-exchange chromatography | (1) Anion-exchange chromatography | ||
(2) Hydrophobic interaction chromatography | (2) Hydrophobic interaction chromatography | ||
(3) Gel-filtration chromatography | (3) Gel-filtration chromatography | ||
'''D) Protein Concentrated''': by use of a Centricon system (Amersham Biosystems). | '''D) Protein Concentrated''': by use of a Centricon system (Amersham Biosystems). | ||
'''E) SDS−PAGE analysis''': of the preparation, detection of protein and contaminants | '''E) SDS−PAGE analysis''': of the preparation, detection of protein and contaminants | ||
'''F) Crystallizations''': Hanging-drop vapor diffusion method | '''F) Crystallizations''': Hanging-drop vapor diffusion method | ||
'''G) Molecular Replacement''': The monoclinic structure was solved via use of the program MOLREP | '''G) Molecular Replacement''': The monoclinic structure was solved via use of the program MOLREP | ||
'''Electron Density aps''': Using [X-Ray Diffraction] were displayed and models were manually constructed with the programs O (33) and XtalView/Xfit | |||
'''Refinement Cycles''': performed with the program CNX, coordinates for the MIO moiety initially taken directly from the HAL structure | '''H) Electron Density aps''': Using [X-Ray Diffraction] were displayed and models were manually constructed with the programs O (33) and XtalView/Xfit | ||
'''I) Refinement Cycles''': performed with the program CNX, coordinates for the MIO moiety initially taken directly from the HAL structure | |||
==References== | ==References== | ||
<references /> | <references /> | ||
