Method of amplifying ATP and use thereof
AssigneeJapan Science and Technology Agency
InventorAkio Kuroda
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Abstract
The ATP amplification method of the present invention is a method for amplifying and detecting a very trace amount of exogenous ATP by allowing a fusion protein (PPK-ADK) of a polyphosphate kinase and an adenylate kinase, the fusion protein not containing ADP, to act on a mixture of ATP, AMP, and a polyphosphate compound. The present invention also provides an ultrasensitive ATP amplification method by which ATP at a single cell level can be amplified and detected, and an ultrasensitive microbial assay based on this ATP amplification method.
Claims3
A method for amplifying adenosine triphosphate comprising:
- allowing a fusion protein that has a polyphosphate kinase and an adenylate kinase in this order from the N-terminal to act on a mixture containing adenosine triphosphate, adenosine monophosphate, and a polyphosphate compound thereby amplifying adenosine triphosphate in the mixture,
- wherein the fusion protein has been subjected to an apyrase treatment and a pyrophosphate treatment so as to remove adenosine diphosphate bound to the fusion protein.
A method for detecting adenosine triphosphate comprising:
- allowing a fusion protein that has a polyphosphate kinase and an adenylate kinase in this order from the N-terminal to act on a mixture of adenosine triphosphate, adenosine monophosphate, and a polyphosphate compound thereby amplifying adenosine triphosphate in the mixture;
- and
- detecting the amplified adenosine triphosphate,
- wherein the fusion protein has been subjected to an apyrase treatment and a pyrophosphate treatment so as to remove adenosine diphosphate bound to the fusion protein.
A method for detecting the presence of a microorganism comprising:
- treating a sample containing a microorganism to prepare a sample containing adenosine triphosphate;
- adding the sample containing adenosine triphosphate to an adenosine triphosphate amplification system thereby amplifying adenosine triphosphate in the sample;
- detecting the amplified adenosine triphosphate;
- and
- detecting the presence of the microorganism by the presence of the detected adenosine triphosphate,
- wherein the adenosine triphosphate amplification system comprises adenosine monophosphate, a polyphosphate compound, and a fusion protein that has a polyphosphate kinase and an adenylate kinase in this order from the N-terminal,
- wherein the fusion protein has been subjected to an apyrase treatment and a pyrophosphate treatment so as to remove adenosine diphosphate bound to the fusion protein.
Dependencies are inferred from the claim text.