Flowable resins used for dental restoration are subject to biofilm formation. Zinc has antibacterial properties. Thus, we prepared a zinc-doped phosphate-based glass (Zn-PBG) to dope a flowable resin and evaluated the antibacterial activity of the composite against Streptococcus mutans (S. mutans) to extrapolate the preventative effect toward secondary caries. The composites were prepared having 0 (control), 1.9, 3.8, and 5.4 wt.% Zn-PBG. The flexural strength, elastic modulus, microhardness, depth of cure, ion release, inhibition zone size, and number of colony-forming units were evaluated and analyzed using ANOVA. The flexural strength of the control was significantly higher than those of Zn-PBG samples (p < 0.05). However, all samples meet the International Standard, ISO 4049. The microhardness was not significantly different for the control group and 1.9 and 3.8 wt.% groups, but the 5.4 wt.% Zn-PBG group had a significantly lower microhardness (p < 0.05). Further, the composite resins increasingly released P, Ca, Na, and Zn ions with an increase in Zn-PBG content (p < 0.05). The colony-forming unit count revealed a significant reduction in S. mutans viability (p < 0.05) with increase in Zn-PBG content. Therefore, the addition of Zn-PBG to flowable composite resins enhances antibacterial activity and could aid the prevention of secondary caries.

Alteromonas sp. GNUM-1 is known to degrade agar, the main cell wall component of red macroalgae, for their growth. A putative agarase gene (agaG1) was identified from the mini-library of GNUM-1, when extracellular agarase activity was detected in a bacterial transformant. The nucleotide sequence revealed that AgaG1 had significant homology to GH16 agarases. agaG1 encodes a primary translation product (34.7 kDa) of 301 amino acids, including a 19-amino-acid signal peptide. For intracellular expression, a gene fragment encoding only the mature form (282 amino acids) was cloned into pGEX-5X-1 in Escherichia coli, where AgaG1 was expressed as a fusion protein with GST attached to its N-terminal (GST-AgaG1). GST-AgaG1 purified on a glutathione sepharose column had an apparent molecular weight of 59 kDa on SDS-PAGE, and this weight matched with the estimated molecular weight (58.7 kDa). The agarase activity of the purified protein was confirmed by the zymogram assay. GST-AgaG1 could hydrolyze the artificial chromogenic substrate, p-nitrophenyl-β-D-galactopyranoside but not p-nitrophenyl-α-D-galactopyranoside. The optimum pH and temperature for GST-AgaG1 activity were identified as 7.0 and 40 °C, respectively. GST-AgaG1 was stable up to 40 °C (100 %), and it retained more than 70 % of its initial activity at 45 °C after heat treatment for 30 min. The K m and V max for agarose were 3.74 mg/ml and 23.8 U/mg, respectively. GST-AgaG1 did not require metal ions for its activity. Thin layer chromatography analysis, mass spectrometry, and (13)C-nuclear magnetic resonance spectrometry of the GST-AgaG1 hydrolysis products revealed that GST-AgaG1 is an endo-type β-agarase that hydrolyzes agarose and neoagarotetraose into neoagarobiose.

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