Expression, purification, crystallization, and preliminary X-ray analysis of the N-terminal domain of Escherichia coli adenylyl transferase.
A soluble N-terminal domain of the Escherichia coli adenylyl transferase (ATase) is responsible for deadenylylation activity of the intact enzyme. Previous studies of the deadenylylation activity have involved a fragment, AT-N423 (residues 1 to 423), which was extended by 17 amino acids to give AT-N440. This new domain is truncated at the end of a predicted helix and prior to a Q-linker. The domain was found to be very soluble and stable so that it could be purified to homogeneity and crystallized. This construct has deadenylylation activity that is independent of the low nitrogen status indicator PII-UMP. The crystals belong to space group P3(1)21 or its enantiomorph P3(2)21 with a=b=116.6 A and c=67.6 A.
Xu
Yibin
Y
Department of Biochemistry and Molecular Biology, James Cook University, Townsville, Queensland 4811, Australia. yibin.xu@jcu.edu.au
Wen
Daying
D
Clancy
Paula
P
Carr
Paul D
PD
Ollis
David L
DL
Vasudevan
Subhash G
SG
D000249:Adenosine Monophosphate / Q000378:metabolism; D000595:Amino Acid Sequence; D000837:Anion Exchange Resins / Q000737:chemistry; D015153:Blotting, Western; D002845:Chromatography / Q000379:methods; D003460:Crystallization; D018360:Crystallography, X-Ray; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D015870:Gene Expression / Q000235:genetics*; D005822:Genetic Vectors / Q000235:genetics; D005974:Glutamate-Ammonia Ligase / Q000378:metabolism; D005971:Glutamates / Q000032:analysis / Q000378:metabolism; D006877:Hydroxamic Acids / Q000032:analysis / Q000378:metabolism; D007656:Ketoglutaric Acids / Q000737:chemistry / Q000378:metabolism; D008969:Molecular Sequence Data; D009713:Nucleotidyltransferases / Q000096:biosynthesis* / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D051922:PII Nitrogen Regulatory Proteins; D010446:Peptide Fragments / Q000096:biosynthesis* / Q000737:chemistry; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000302:isolation & purification / Q000378:metabolism; D012685:Sepharose / Q000031:analogs & derivatives* / Q000737:chemistry
D000837:Anion Exchange Resins; D005971:Glutamates; D006877:Hydroxamic Acids; D007656:Ketoglutaric Acids; D051922:PII Nitrogen Regulatory Proteins; D010446:Peptide Fragments; D011994:Recombinant Proteins; C050614:glutamate-gamma-hydroxamic acid; D000249:Adenosine Monophosphate; C020927:phenyl-sepharose; D012685:Sepharose; D009713:Nucleotidyltransferases; C020658:glutamine-synthetase adenylyltransferase; C022515:regulatory protein uridylyltransferase; D005974:Glutamate-Ammonia Ligase