Different Physiological Roles of ATP- and PPi-Dependent Phosphofructokinase Isoenzymes in the Methylotrophic ActinomyceteAmycolatopsis methanolica

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Abstract
Cells of the actinomyceteAmycolatopsis methanolicagrown on glucose possess only a single, exclusively PPi-dependent phosphofructokinase (PPi-PFK) (A. M. C. R. Alves, G. J. W. Euverink, H. J. Hektor, J. van der Vlag, W. Vrijbloed, D.H.A. Hondmann, J. Visser, and L. Dijkhuizen, J. Bacteriol. 176:6827–6835, 1994). When this methylotrophic bacterium is grown on one-carbon (C1) compounds (e.g., methanol), an ATP-dependent phosphofructokinase (ATP-PFK) activity is specifically induced, completely replacing the PPi-PFK. The twoA. methanolicaPFK isoenzymes have very distinct functions, namely, in the metabolism of C6and C1carbon substrates. This is the first report providing biochemical evidence for the presence and physiological roles of PPi-PFK and ATP-PFK isoenzymes in a bacterium. The novel ATP-PFK enzyme was purified to homogeneity and characterized in detail at the biochemical and molecular levels. TheA. methanolicaATP-PFK and PPi-PFK proteins possess a low level of amino acid sequence similarity (24%), clearly showing that the two proteins are not the result of a gene duplication event. PPi-PFK is closely related to other (putative) actinomycete PFK enzymes. Surprisingly, theA. methanolicaATP-PFK is most similar to ATP-PFK from the protozoonTrypanosoma bruceiand PPi-PFK proteins from the bacteriaBorrelia burgdorferiandTreponema pallidum, both spirochetes, very distinct from actinomycetes. The data thus suggest thatA. methanolicaobtained the ATP-PFK-encoding gene via a lateral gene transfer event.