Chapter 11 : Carbohydrate Catabolism: Pathways, Enzymes, Genetic Regulation, and Evolution

Catabolism of hexoses, linear hexitols, and sucrose in B. subtilis. Transport and enzymatic steps are symbolized by arrows. The names of the structural genes (in some cases putative) of the relevant proteins are indicated in italics (lev = levD, levE, levF, and levG). All these genes are both mapped (except pgi, for phosphoglucoisomerase) and inducible (except fruC, pfk, and pgi). Various controls prevent (or reduce) simultaneous expression of alternative pathways for sucrose and fructose (see text). Transfructosylation from sucrose by Lvs (sacB) results mainly in a mixture of glucose, fructose, and levan. sacC encodes levanase. Fra, Glu, Gut, Md, and Suc: fructose, glucose, glucitol, mannitol, and sucrose, respectively; Suc-P, sucrose-phosphate (phosphorylated on position 6 of glucose moiety). Other sugar-phosphates are designated according to standard conventions; for example, Glu-6-P and Fru-l,6-dP designate glucose 6-phosphate and fructose 1,6-diphosphate.

Organization of the gluconate (gnt) operon and the xyloside-xylose (xyn-xyl) regulon in B. subtilis. The repressor of the gnt operon is encoded by the first gene (gntR) of the operon; gntK, -P, and -Z very likely encode gluconate kinase, gluconate permease, and 6-phosphogluconate dehydrogenase, respectively; gntZ is transcribed from both the GntR-repressed promoter and a constitutive promoter within gntP. The repressor of the xylose-xyloside regulon (XylR) controls transcription of the xynCB and xylAB opérons; XynB is a xylosidase; XynC appears to be a membrane protein that might be a permease of xyloside and/or xylose; and xylA and xylB encode xylose isomerase and xylulose kinase, respectively. t, terminator.

Regulons dependent on antiterminators and/or PTS control: comparison of regulation and genetic organization. The first three systems (a, b, and c) involve elements both structurally and functionally homologous: antiterminators, conditional terminators (ct), and enzymes II. (a) E. coli β-glucoside (bgl) operon model. In the absence of β-glucoside inducer, transcription of the operon from the promoter is prevented by two conditional terminators bracketing the antiterminator bglG gene. In these conditions, BglG function is inhibited by phosphorylation by the bglF gene product (enzyme IIβ-glucoside). The presence of β-glucoside in the medium results in its transport and phosphorylation by enzyme IIβ-glucoside. This results in phosphate group rerouting. Dephosphorylated antiterminator then allows full transcription of the operon. BglB is the phospho-β-glucosidase ( 3 , 91 ). (b) B. subtilis sacB (Lvs) regulon. This regulon comprises the sacB gene and the unlinked sacXY regulator operon. Both appear to be controlled by means of a regulatory cascade similar to that of the bgl operon, i.e., positively by both sucrose and SacY antiterminator and negatively by SacX, a putative enzyme IIsucrose. Several regulatory cross-talks with the sacPA-sacT regulon (see below) are not shown. The deg genes (DEG; see chapter 50) activate transcription from both sacB and sacXY promoters ( 13 , 14 , 98 , 104 ). (c) B. subtilis sacPA operon. Transcription of this operon is positively controlled by sucrose and the antiterminator encoded by the linked sacT gene; orf-2 appears not to play a role in this regulation ( 15 ). The mechanism of SacT activation by the presence of sucrose is unknown (see text), (d) B. subtilis levanase operon. The first four genes encode an enzyme IIfructose (very poorly related to those mentioned above) controlling the LevR positive regulator. The fructose-dependent induction cascade appears similar to that of the bgl operon but appears not to involve antitermination. LevR is a transcriptional activator binding upstream from the operon promoter and requiring σ54 as a cofactor ( 16 , 17 , 67 ). (e) B. subtilis glycerol regulon. This regulon comprises at least two unlinked gene clusters and three promoters likely controlled by the glpP gene product. This regulator (unrelated to BglG) appears to function as an antiterminator at conditional terminators just upstream from glpD and glpFK ( 48 – 50 ).

Sucrose gene clusters in four bacteria. The comparison includes two gram-positive systems, from B. subtilis and S. mutans, and two gram-negative systems (sucrose operon in V. alginolyticus, a close relative of enteric bacteria, and sucrose genes present on the S. typhimurium pUR400 plasmid; the pUR400 sucrose regulon appears to be closely related to the chromosomal K. pneumoniae sucrose system). The four enzymes nsucrose, encoded by sacP (in B. subtilis) or scrA genes, are structurally homologous. The S. mutatis ScrA protein contains a C-terminal extension containing a second phosphorylation site (enzyme IIIsucrose domain). The gene designated sacA in B. subtilis or scrB in S. mutons and V. alginolyticus encodes related phosphosucrases (the pUR400 scrB gene sequence is unknown). Expression of the S. mutans sucrose genes is stimulated by the presence of different sugars in the medium (see text). The three other systems are specifically induced by sucrose via activation of the SacT antiterminator in B. subtilis or inactivation of ScrR repressors in the case of the pUR400 or V. alginolyticus systems. The two ScrR repressors are not strongly similar, but both belong to the LacI-GalR superfamily; the pUR400-encoded ScrR repressor binds fructose; the two gram-negative regulons are dependent on the cyclic-AMP receptor protein-cyclic-AMP complex. The scrK and scrY genes encode fructokinase and a sucrose porin (related to the LamB maltoporin), respectively, orf encodes unknown functions (probably not involved in sucrose metabolism) (see references 7 , 15 , 87 , and 89 and references therein). The S. xylosus sucrose genes were recendy cloned and sequenced; the gene products are homologous to those mentioned above. scrA (encoding a short enzyme II) and scrB are unlinked. Both are sucrose inducible via a negative control. Similar operatorlike sequences are found in both leader regions ( 114a ).

FTS-dependent lactose systems of S. aureus and some lactic acid-producing bacteria: pathways and gene organization. (A) Transport and catabolism of lactose via the tagatose 6-phosphate pathway. Lactose is transported and phosphorylated by enzyme IILac (IIlac) and then hydrolyzed into glucose (catabolized by the classical glycolysis pathway) and Gal-6-P, which is isomerized to tagatose 6-phosphate (Tag-6-P). Tag-6-P is converted, by means of a pathway similar to that of its isomer, fructose 6-phosphate (Fru-6-P), into two molecules of triose-phosphate (dihydroxyacetone-phosphate [DHA-P] and glyceraldehyde 3-phosphate [GA-3-P]) and then into phosphoenolpyru-vate (PEP). Dephosphorylation of PEP to pyruvate results partly in ATP production (not shown) and partly in phosphorylation of proteins of the cytoplasmic PTS cascade. This cascade ends with phosphorylation of the membrane enzyme IILac protein by the cytoplasmic enzyme IIILac (IIIlac) protein. In lactic acid bacteria, lactate produced from pyruvate is excreted with protons; this-coexcretion contributes to energizing the proton motive force. A, H, I, and K, aldolases, hydrolases, isomerases, and kinases, respectively. The genes encoding the enzymes of the pathway are indicated in parentheses. (B) Organization of the lac opérons in L. lactis, S. aureus, and L. casei. Arrows indicate the transcripts; t, t1, and t2 are transcriptional terminators. Three major features distinguish the L. lactis lac operon from that of S. aureus: the orientation of the lacR gene, the presence of a weak terminator (tl) downstream from lacE, and the presence of an eighth gene, lacX of unknown function, at the 3′ end of the operon. The 5′ end of the L. casei operon has not yet been sequenced, but its transcript is similar in size to that of S. aureus. (Adapted from references 2 , 19 , 72 , and 114. )