Cyclic AMP affinity purification and ESI–QTOF MS–MS identification of cytosolic glyceraldehyde 3‐phosphate dehydrogenase and two nucleoside diphosphate kinase isoforms from tobacco BY‐2 cells

[摘要]

The soluble protein fraction of tobacco bright yellow 2 cells contained adenosine 3′,5′-cyclic monophosphate (cAMP)-binding activity, detected with both a conventional binding assay and a surface plasmon resonance biosensor. A cAMP-agarose-based affinity purification procedure yielded three proteins which were identified by mass spectrometry as glyceraldehyde 3-phosphate dehydrogenase (GAPDH) and two nucleoside diphosphate kinases (NDPKs). This is the first report describing an interaction between cAMP and these proteins in higher plants. Our findings are discussed in view of the reported role of the interaction of cAMP with GAPDH and NDPK in animals and yeast. In addition, we provide a rapid method to isolate both proteins from higher plants.

[学科分类] 生物化学/生物物理

[关键词] Adenosine 3′;5′-cyclic monophosphate;Cell cycle;Glyceraldehyde 3-phosphate dehydrogenase;Nucleoside diphosphate kinase;Tobacco bright yellow 2;AET-cAMP;8-(2-aminoethylthio)-cAMP;BSA;bovine serum albumin;cAMP;adenosine 3′;5′-cyclic monophosphate;ESI;electrospray ionisation;GAPDH;glyceraldehyde 3-phosphate dehydrogenase;MS;mass spectrometry;NAD+;β-nicotinamide adenine dinucleotide;NDPK;nucleoside diphosphate kinase;PKA;protein kinase A;TBY-2;tobacco bright yellow 2