Involvement of Phosphoenolpyruvate in the Catabolism of Caries-Conducive Dĭsaccharides by Streptococcus mutans : Lactose Transport

Abstract

The mechanisms for transport and hydrolysis of lactose were investigated in five cariogenic strains (HS6, AHT, FA1, NCTC 10449, and SL1) representing the four serogenetic groups of Streptococcus mutans. The systems for transport and hydrolysis of lactose had the characteristics of a phosphoenolpyruvate (PEP)-dependent lactose (Lac) phosphotransferase (PT) system and phospho-β-galactosidase (P-β-gal), respectively, in all strains tested, except strain HS6. Decryptified cells required PEP and Mg 2+ for transport of the non-metabolizable model β-galactosides o -nitrophenyl-β- d -galactopyranoside (ONPG) and thiomethyl-β- d -galactopyranoside (TMG). Substitution of 2-phosphoglycerate (2-PG) for PEP also stimulated the Lac PT system. Other potential high-energy phosphate donors (adenosine tri-, di-, and monophosphates and guanosine triphosphate) did not stimulate the Lac PT system. Sodium fluoride had no effect upon the PEP-dependent Lac PT system in decryptified cells with PEP as the energy source; however, when 2-PG was used as the energy source, F − inhibited ONPG phosphorylation. With intact cells which must generate PEP endogenously, the presence of F − in concentration ≥ 10 mM completely inhibited the Lac PT system, presumably through inhibition of 2-PG hydrolyase (EC 4.2.1.11; enolase). Both intact and decryptified cells accumulated a phosphorylated derivative of TMG that behaved chromatographically as TMG-phosphate. After alkaline phosphatase treatment, the derivative had an R f identical to that of TMG. No β-galactosidase (β-gal) activity was detected with ONPG as the substrate; hydrolysis occurred only when ONPG-6-phosphate was supplied as the substrate. Strain HS6 apparently transported lactose by an active transport-type system in which the accumulated intracellular product was the free disaccharide based on the following criteria: (i) ONPG transport and hydrolysis in decryptified cells was not stimulated by PEP; (ii) ONPG hydrolysis occurred in the absence of PEP; and (iii) ONPG-6-phosphate was not hydrolyzed. These data indicate that, in all strains tested except strain HS6, lactose transport was mediated by a PEP-dependent Lac PT system, resulting in accumulation of lactose-phosphate that was hydrolyzed by an enzyme similar to the P-β-gal of group N streptococci and Staphylococcus aureus; conversely, strain HS6 transported and hydrolyzed lactose by a PEP-independent transport system and β-gal, respectively.