5). In UA159, cystine starvation resulted in Selleckchem AZD6738 a lower growth yield as well as a longer doubling time (Tdc. 93.3 ± 0.7 min) compared with its growth in the presence of cystine (Tdc. 76.3 ± 1.5 min), indicating that l-cystine is required for optimal growth of S. mutans. However, growth was completely abolished in SmTycABC under cystine starvation. Supplementing the modified growth medium with 0.1 mM cystine slightly improved the drastic growth impairment of the SmTcyABC mutant (Tdc. 118.2 ± 0.8 min). Similar to the SmTcyABC transporter mutant, the TcyR-deficient mutant (SmTcyR) had a longer doubling time (Tdc. 117.2 ± 3.8 min)
under cystine-supplemented (1 mM) conditions relative to wild type (Fig. 5). In contrast to SmTcyABC, SmTcyR was able to survive under cystine-deficient conditions, although its doubling time was remarkably increased relative to wild type (Tdc. 261.0 ± 11.9 min). Also importantly, growth kinetics of SmTcyR revealed a notable increase in the lag time regardless of the presence or absence of cystine, compared with the wild-type UA159 and SmTcyABC. We further evaluated the effect on growth by individual components of the TcyABC operon by conducting growth studies on mutants deficient in each gene. Briefly, growth kinetics were monitored for the TcyA, this website TcyB, and TcyC
transporter mutants in modified MM without cystine (Fig. 6). The most drastic effect on growth was observed for SmTcyB. Similar to TcyABC, growth of this mutant was completely abolished without cystine. Although TcyA and TcyC were able to grow in cystine-deficient medium, their
growth was tremendously impaired relative to wild type as judged by their longer doubling times; Tdc. 131.3 ± 4.8 and 214.8 ± 21.5 min, respectively. Sperandio et al. 2010 also showed impaired growth in the form of pinpoint colonies when their TcyA mutant was grown in chemically defined medium with the addition of cystine as the sole sulfur source. However, they did not investigate the growth of other Tyc ABC mutants. The ability of some of our TycABC mutants to grow in the absence of cystine, albeit in an impaired fashion, suggests that the presence of other amino acids (i.e. glutamate and leucine), inorganic sulfur, and/or ammonium sources were sufficient to sustain growth. S. mutans possesses amino acid biosynthetic pathways and even though most amino acids are not freely available in the Acyl CoA dehydrogenase environment, some strains are able to synthesize all the necessary amino acids required for survival (Liu & Ferro-Luzzi Ames, 1998; Albanesi et al., 2005). The ability of S. mutans to scavenge and compete for limited nutrients in the plaque biofilm is an important aspect that confers an ecological advantage, which facilitates its survival and persistence in the oral cavity. The amino acid transport system in S. mutans UA159, encoded by the tcyABC operon that is induced under cystine-starved conditions, functions to maintain growth by transporting cystine into the cell.