ATP-reactive sites in the bacteriophage lambda packaging protein terminase lie in the N-termini of its subunits, gpA and gpNu1.
ATP-reactive sites in terminase and its subunits have been successfully identified using three different affinity analogs of ATP (2-and 8-azidoATP and FITC) GpA, the larger subunit of terminase, was shown to have a higher affinity for these analogs than gpNu1, the smaller subunit. The suitability of these reagents as affinity analogs of ATP was demonstrated by ATP protection experiments and in vitro assays done with the modified proteins. These analogs were thus shown to modify the ATP-reactive sites. The results obtained from these experiments also indicate the importance of subunit-subunit interactions in the holoenzyme. Terminase, gpA, and gpNu1 were modified with these analogs and the ATP-reactive sites were identified by isolating the modified peptide by reverse-phase chromatography. The sequence analysis of the modified peptides indicates a region including amino acids 18-35 in the N-terminus of gpNu1 and a region including amino acids 59-85 in the N-terminus of gpA as being the ATP-reactive sites.
Department of Molecular and Medical Genetics, University of Toronto, Canada.
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D000251:Adenosine Triphosphatases / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000378:metabolism*; D000345:Affinity Labels / Q000378:metabolism; D001386:Azides / Q000378:metabolism; D010582:Bacteriophage lambda / Q000187:drug effects* / Q000201:enzymology; D001665:Binding Sites; D002845:Chromatography; D004706:Endodeoxyribonucleases / Q000302:isolation & purification / Q000378:metabolism*; D004926:Escherichia coli / Q000821:virology; D016650:Fluorescein-5-isothiocyanate / Q000378:metabolism; D010455:Peptides / Q000302:isolation & purification; D019920:Photoaffinity Labels / Q000378:metabolism; D019065:Virus Assembly
D000345:Affinity Labels; D001386:Azides; D010455:Peptides; D019920:Photoaffinity Labels; C017040:8-azidoadenosine 5'-triphosphate; D000255:Adenosine Triphosphate; D004706:Endodeoxyribonucleases; C030894:terminase; D000251:Adenosine Triphosphatases; D016650:Fluorescein-5-isothiocyanate