Disruption of the adh (acetoin dehydrogenase) operon has wide-ranging effects on Streptococcus mutans growth and stress response

The agent largely responsible for initiating dental caries, Streptococcus mutans produces acetoin dehydrogenase that is encoded by the adh operon. The operon consists of the adhA and B genes (E1 dehydrogenase), adhC (E2 lipoylated transacetylase), adhD (E3 dihydrolipoamide dehydrogenase), and lplA (lipoyl ligase). Evidence is presented that AdhC interacts with SpxA2, a redox-sensitive transcription factor functioning in cell wall and oxidative stress responses. In-frame deletion mutations of adh genes conferred oxygen-dependent sensitivity to slightly alkaline pH (pH 7.2-7.6), within the range of values observed in human saliva. Growth defects were also observed when glucose or sucrose served as major carbon sources. A deletion of the adhC orthologous gene, acoC gene of Streptococcus gordonii , did not result in pH sensitivity or defective growth in glucose and sucrose. The defects observed in adh mutants were partially reversed by addition of pyruvate. Unlike most 2-oxoacid dehydrogenases, the E3 AdhD subunit bears an N-terminal lipoylation domain nearly identical to that of E2 AdhC. Changing the lipoyl domains of AdhC and AdhD by replacing the lipoate attachment residue, lysine to arginine, caused no significant reduction in pH sensitivity but the adhDK43R mutation eliminating the lipoylation site resulted in an observable growth defect in glucose medium. The adh mutations were partially suppressed by a deletion of rex , encoding an NAD + /NADH-sensing transcription factor that represses genes functioning in fermentation. spxA2 adh double mutants show synthetic growth restriction at elevated pH and upon ampicillin treatment. These results suggest a role for Adh in stress management in S. mutans . IMPORTANCE Dental caries is often initiated by Streptococcus mutans , which establishes a biofilm and a low pH environment on tooth enamel surfaces. The current study has uncovered vulnerabilities of S. mutans mutant strains that are unable to produce the enzyme complex, acetoin dehydrogenase (Adh). Such mutants are sensitive to modest increases in pH to 7.2-7.6, within the range of human saliva, while a mutant of a commensal Streptococcal species is resistant. The S. mutans adh strains are also defective in carbohydrate utilization and are hypersensitive to a cell wall-acting antibiotic. The studies suggest that Adh could be a potential target for interfering with S. mutans colonization of the oral environment.

An Extended Gate Field Effect Transistor (EGFET) pH Microsensor

Here, an extended – gate field effect transistor (EGFET) pH microsensor was developed for use in fast and sensitive pH measurement applications. The system consisted of two components; a pH sensitive modified gold electrode and a simple and cheap metal oxide semiconductor field-effect transistor (MOSFET). Polypyrrole, a semiconductor and pH responsive polymer, was formed by electropolymerization of pyrrole monomer at the surface of the gold electrode in galvostanic mode. Then, measurements were made in PBS at different pH values using the pH sensitive electrode. In this context, the pH sensitivity of polypyrrole with respect to electropolymerization and incubation time were studied. According to the results, the EGFET pH microsensor formed by 4-min pyrrole electropolymerization showed at pH 6-12 the highest pH sensitivity with 67 mV/pH

An Extended Gate Field Effect Transistor (EGFET) pH Microsensor

Here, an extended – gate field effect transistor (EGFET) pH microsensor was developed for use in fast and sensitive pH measurement applications. The system consisted of two components; a pH sensitive modified gold electrode and a simple and cheap metal oxide semiconductor field-effect transistor (MOSFET). Polypyrrole, a semiconductor and pH responsive polymer, was formed by electropolymerization of pyrrole monomer at the surface of the gold electrode in galvostanic mode. Then, measurements were made in PBS at different pH values using the pH sensitive electrode. In this context, the pH sensitivity of polypyrrole with respect to electropolymerization and incubation time were studied. According to the results, the EGFET pH microsensor formed by 4-min pyrrole electropolymerization showed at pH 6-12 the highest pH sensitivity with 67 mV/pH

Fabrication And pH Sensing Characteristics Measurement of Back Gate ZnO Thin Film Planar FET

Here in we demonstrate the design of a low- cost zinc oxide (ZnO) thin-film planar transistor-based pH sensor controlled by the bottom gate fabricated by a fairly simple fabrication approach. The performance of the fabricated device is evaluated by electrical as well as surface characterization. The surface morphology is analyzed by scanning electron microscope (SEM) and atomic force microscopy (AFM) and it shows surface properties that are essential for a device to function as a pH sensor. The fabricated thin-film FET comprises Zinc Oxide (ZnO) as a channel layer of length 6 µm and thickness 200 nm, Silicon Nitride (Si3N4) as a pas- sivation layer, and Aluminum (Al) as a contact layer. The effect on pH sensitivity for varied channel lengths (6 µm, 12 µm, and 15 µm) is also examined and opti- mum results have been achieved at channel length = 6 µm. The change in threshold voltage (ΔVth) & change in current (ΔImax) are used as a sensing metrics to an- alyze the sensing performance of the device. The device shows excellent pH sensitivity in terms of average cur- rent and average voltage sensitivity 120.97 mA/pH and 97.85 mv/pH respectively at pH ranging from 3.2 to 11.1 with best pH stability (linearity) for pH value 4 to 10. The voltage sensitivity is higher than the Nernstian value (59 mv/pH) at room temperature.

Structure and mechanism of the human NHE1-CHP1 complex

Sodium/proton exchanger 1 (NHE1) is an electroneutral secondary active transporter present on the plasma membrane of most mammalian cells and plays critical roles in regulating intracellular pH and volume homeostasis. Calcineurin B-homologous protein 1 (CHP1) is an obligate binding partner that promotes NHE1 biosynthetic maturation, cell surface expression and pH-sensitivity. Dysfunctions of either protein are associated with neurological disorders. Here, we elucidate structures of the human NHE1-CHP1 complex in both inward- and inhibitor (cariporide)-bound outward-facing conformations. We find that NHE1 assembles as a symmetrical homodimer, with each subunit undergoing an elevator-like conformational change during cation exchange. The cryo-EM map reveals the binding site for the NHE1 inhibitor cariporide, illustrating how inhibitors block transport activity. The CHP1 molecule differentially associates with these two conformational states of each NHE1 monomer, and this association difference probably underlies the regulation of NHE1 pH-sensitivity by CHP1.