## 1 Introduction  
Glucose metabolism in most bacteria starts via its uptake and phosphorylation (either by a PTS or via glucokinase) to glucose 6-phosphate (G6P) (for reviews: [1,2]). G6P is subsequently broken down to pyruvate mainly via the Embden-Meyerhof-Parnas pathway (EMP), and in part by the hexose monophosphate shunt (HMP), or the Entner-Doudoroff (ED) pathway [3]. In the EMP, G6P is isomerized by phosphoglucose isomerase (PGI) to fructose 6-phosphate (F6P). The first committed step in EMP glycolysis is catalyzed by phosphofructokinase (fructose 6-phosphate 1-kinase, PFK) which phosphorylates F6P to fructose 1,6-bisphosphate (F1,6 BP) in a practically irreversible reaction [4,5]. PFK thus plays a pivotal role in the regulation of and flux through the EMP pathway of glycolysis. Furthermore, the oxidative pentose phosphate pathway (PPP) also depends on PFK activity. The model microorganism, _Escherichia coli_ (_E. coli_) K-12 has two genes encoding PFK activity (_p/fkA, p/fkB_) [6,7]; the encoded isozymes are quite different as they display distinct kinetic features and they are differently regulated [4,8]. Mutations which led to the loss of the major activity (PFKA, alternatively abbreviated in the literature as PFK-1) already yielded growth defects on carbon sources such as glucose or hexitols (mannitol, glucitol/sorbitol) [6,7]. _E. coli pfKA_ mutants gave rise to suppressor mutations, (e.g., _pfkB_1) with elevated enzyme activity of PFK-2 (PFKB) [6,7,9]. Double deletions (\(\Delta\)_pfKA_, \(\Delta\)_pfkB_) are devoid of PFK activity and showed severe growth defects on several sugars such as glucose [10, 11, 12, 13]. Growth of _pfk_ mutants on fructose, however, is less affected as this hexose mainly enters via the fructose PTS (EII\({}^{\text{Fru}}\)) which phosphorylates it to fructose 1-phosphate which is further converted by a fructose 1-phosphate 6-kinase [14]. In _pfk_ mutants, G6P may be channeled via glucose 6-phosphate dehydrogenase (ZWF) to the oxidative pentose phosphate pathway (PPP) and/or to the Entner-Doudoroff (ED) pathway of glycolysis [6,7]. From the PPP, glyceraldehyde 3-phosphate (G3P) could enter the lower glycolytic trunk [15]. A deletion of the _zwf_ gene would block both the entry to the oxidative PPP and to the ED pathway. Recently, three groups have reported the construction of _E. coli_ triple deletion strains (\(\Delta\)_pfkA_, \(\Delta\)_pfkB_, \(\Delta\)_zwf_) but did not describe growth behavior on glucose [16, 17, 18]. A \(\Delta\)_zwf_ \(\Delta\)_pfkB_ \(\
ot\)_pA_::114-_pfkA_ (DAS + 4) \"PFK-knockdown\" mutant of strain MG1655 has been used for redirecting flux from G6P to inositol formation [19]. Such a triple-negative mutant would be expected to be devoid of growth on glucose and several other C sources (Figure 1). Our group has discovered two genes (_fsaA_, _fsaB_) in the _E. coli_ K-12 chromosome which both encode fructose 6-phosphate aldolase (FSA) activity (forming dihydroxyacetone (DHA) and G3P from F6P in a reversible manner) when expressed from recombinant vectors [20]. Both genes, however, are expressed at very low rates from their chromosomal locations ([21,22], and our own results); estimations for a single _E. coli_ cell (after growth in minimal media) gave values in the range of 100 protein molecules of FSAA (gene product of _fsaA_ gene) or even less for FSAB [22]. The physiological roles (in vivo) of the two genes therefore remain enigmatic [23]. Recombinant FSAA as an in vitro aldolase biocatalyst, however, has been the subject of many investigations in chemo-enzymatic syntheses as FSAA displays an unusual broad substrate spectrum encompassing not only DHA but also other donor compounds such as hydroxyacetone (acetol) or glycolaldehyde [24, 25, 26, 27]. For utilization of FSA activity in vivo, several groups have successfully utilized the cloned _fsaA_ gene for metabolic pathway engineering of _E. coli_. By doing so, growth on DHA [28], growth on  
Figure 1: Schematic of the glycolytic blockage in the triple deletion strain, GL3.