Tobacco etch virus (TEV) protease is a 27‐kDa catalytic domain of the polyprotein nuclear inclusion a (NIa) in TEV, which recognizes the specific amino acid sequence ENLYFQG/S and cleaves between Q and G/S. Despite its substrate specificity, its use is limited by its autoinactivation through self‐cleavage and poor solubility during purification. It was previously reported that T17S/N68D/I77V mutations improve the solubility and yield of TEV protease and S219 mutations provide protection against self‐cleavage. In this study, we isolated TEV proteases with S219N and S219V mutations in the background of T17S, N68D, and I77V without the inclusion body, and measured their enzyme kinetics. The k _cat of two isolated S219N and S219V mutants in the background of T17S, N68D, and I77V mutations was highly increased compared to that of the control, and S219N was twofold faster than S219V without K _m change. This result indicates that combination of these mutations can further enhance TEV activity.

Persistent organic pollutants (POPs) are lipid-soluble toxins that are not easily degraded; therefore, they accumulate in the environment and the human body. Several studies have indicated a correlation between POPs and metabolic diseases; however, their effects on mitochondria as a central organelle in cellular metabolism and the usage of mitochondria as functional markers for metabolic disease are barely understood. In this study, a zebrafish model system was exposed to two subclasses of POPs, organochlorine pesticides (OCPs) and polychlorinated biphenyls (PCBs), under two different conditions (solitary OCPs or OCPs with PCBs (Aroclor 1254)), and changes in the oxidative stress marker levels and mitochondrial enzyme activities in the electron transport chain of the tail were measured to observe the correlation between POPs and representative biomarkers for metabolic disease. The results indicated different responses upon exposure to OCPs and OCPs with Aroclor 1254, and accelerated toxicity was observed following exposure to mixed POPs (OCPs with Aroclor 1254). Males were more sensitive to changes in the levels of oxidative stress markers induced by POP exposure, whereas females were more susceptible to the toxic effects of POPs on the levels of mitochondrial activity markers. These results demonstrate that the study reflects real environmental conditions, with low-dose and multiple-toxin exposure for a long period, and that POPs alter major mitochondrial enzymes’ functions with an imbalance of redox homeostasis in a sex-dependent manner.

Type 2 diabetes (T2D) is characterized by defects in insulin action to target tissues, resulting in hyperglycemia, insulin resistance, and mitochondrial dysfunction. The eye is one of the organs susceptible to T2D, but knowledge regarding mitochondrial dysfunction in the eyes after hyperglycemia and T2D is based mainly on epidemiological evidence, with little experimental data. POPs are known as endocrine-disrupting chemicals and are associated with uncontrolled glucose and lipid metabolism, leading to the onset of diabetes. To determine the relationship between POPs and T2D, two model systems were developed: glucose-immersed zebrafish to induce hyperglycemia, and zebrafish exposed to low-dose POPs in a water circulating system for three months. To examine the role of mitochondrial function, the activity of mitochondrial complexes I, II, III, and IV from the eyes of the two zebrafish models was measured spectrophotometrically. Enhanced enzymatic activities of mitochondrial complexes III and IV were observed in the eyes of both hyperglycemia and low-dose POPs exposed models, especially in male zebrafish. These results demonstrate that POPs alleviate mitochondrial oxidative phosphorylation (OXPHOS) in a sex-dependent manner through a compensatory mechanism, which is also observed in acute hyperglycemia.

Benzalkonium chloride (BKC) is a cationic surfactant used as a component in ophthalmic eye drops. The effects of BKC on the eye and the simple binding of BKC on the contact lens surface have been reported in other studies. However, the exact value of the BKC binding affinity on the contact lens surfaces and its effects on the physical properties of contact lenses have not been studied. Here, the binding affinity of BKC toward two types of contact lenses, those with the wetting agent polyvinylpyrrolidone (PVP) and those without, was calculated. In addition, the refractive power, UV-vis transmittance, contact angle, water content, base curve, and diameter of the contact lenses after treatment with BKC were examined to evaluate the effects of its adsorption on the contact lens. We found that the maximum amount of adsorbed BKC was 2.88 mM for the contact lens without PVP whereas it was 2.32 mM for that with PVP. In contrast, the BKC binding affinities were similar. Crucially, the physical properties of the contact lens changed significantly because of the adsorption of BKC. Although BKC is a widely used preservative, our results suggest that use of PVP-containing contact lenses reduces BKC adsorption and discomfort.

The degree of protonation of contact lens materials is affected by the surrounding pH environment, due to the different pK_a values. The swelling of ionic contact lenses is generally controlled by these factors which determines physical properties of contact lenses. The purpose of this study was to evaluate the pH dependence of the physical properties of contact lenses. The ionic etafilcon A and non-ionic hilafilcon B contact lenses were used in this study. The diameter, refractive power, equilibrium water content (EWC), and the amounts of freezable-free water (W_ff), freezable-bound water (W_fb) and non-freezable water (W_nf) in the contact lens at each pH condition were measured. The diameter, refractive power and EWC of etafilcon A decreased with decreasing pH below 7.0 or 7.4, whereas hilafilcon B showed relatively constant values. The quantity of W_fb tended to increase with increasing pH, showing a relatively constant value above 7.0, whereas W_nf decreased. Hilafilcon B did not show changes in EWC and specific trends in W_fb and W_nf. The significant change of etafilcon A at more acidic condition is derived from the presence of methacrylic acid (MA) which makes it vulnerable to pH. Additionally, though the EWC is composed of various states of water, (i) various states of water could response to surrounding environment in different way with EWC and (ii) W_fb could be the crucial factor that determines physical properties of contact lens.