Increasing stability of water-soluble PQQ glucose dehydrogenase by increasing hydrophobic interaction at dimeric interface

BACKGROUND: Water-soluble quinoprotein glucose dehydrogenase (PQQGDH-B) from Acinetobacter calcoaceticus has a great potential for application as a glucose sensor constituent. Because this enzyme shows no activity in its monomeric form, correct quaternary structure is essential for the formation of active enzyme. We have previously reported on the increasing of the stability of PQQGDH-B by preventing the subunit dissociation. Previous studies were based on decreasing the entropy of quaternary structure dissociation but not on increasing the interaction between the two subunits. We therefore attempted to introduce a hydrophobic interaction in the dimeric interface to increase the stability of PQQGDH-B. RESULTS: Amino acid residues Asn340 and Tyr418 face each other at the dimer interface of PQQGDH-B, however no interaction exists between their side chains. We simultaneously substituted Asn340 to Phe and Tyr418 to Phe or Ile, to create the two mutants Asn340Phe/Tyr418Phe and Asn340Phe/Tyr418Ile. Furthermore, residues Leu280, Val282 and Val342 form a hydrophobic region that faces, on the other subunit, residues Thr416 and Thr417, again without any specific interaction. We simultaneously substituted Thr416 and Thr417 to Val, to create the mutant Thr416Val/Thr417Val. The temperatures resulting in lose of half of the initial activity of the constructed mutants were increased by 3–4°C higher over wild type. All mutants showed 2-fold higher thermal stability at 55°C than the wild-type enzyme, without decreasing their catalytic activities. From the 3D models of all the mutant enzymes, the predicted binding energies were found to be significantly greater that in the wild-type enzyme, consistent with the increases in thermal stabilities. CONCLUSIONS: We have achieved via site-directed mutagenesis the improvement of the thermal stability of PQQGDH-B by increasing the dimer interface interaction. Through rational design based on the quaternary structure of the enzyme, we selected residues located at the dimer interface that do not contribute to the intersubunit interaction. By substituting these residues to hydrophobic ones, the thermal stability of PQQGDH-B was increased without decreasing its catalytic activity

A New Disorder in UV-Induced Skin Cancer with Defective DNA Repair Distinct from Xeroderma Pigmentosum or Cockayne Syndrome

We report the characterization of a Japanese woman who exhibited many freckles and skin cancers in sun-exposed areas, but displayed no photosensitivity. Fibroblasts (KPSX7) derived from this patient showed similar UV sensitivity to that of normal human fibroblasts. The KPSX7 cells showed normal levels of unscheduled DNA synthesis, recovery of RNA synthesis, recovery of replicative DNA synthesis, protein-binding ability to UV-damaged DNA, and post-translational modification of xeroderma pigmentosum (XP) C. These results indicate that the patient had neither XP nor Cockayne syndrome. Although these results suggest that the KPSX7 cells were proficient in nucleotide excision repair activity, host-cell reactivation (HCR) activity of KPSX7 cells was reduced. Furthermore, introduction of UV damage endonuclease into the cells restored repair activity in the HCR assay to almost normal levels. These results indicate that KPSX7 cells are defective for some types of repair activity in UV-damaged DNA. In summary, the patient had a previously unknown disorder related to UV-induced carcinogenesis, with defective DNA repair

Cerebral cortical processing time is elongated in human brain evolution

サルより遅いヒトの脳処理 --進化するほど脳の回転は遅くなる!?--. 京都大学プレスリリース. 2022-01-26.An increase in number of neurons is presumed to underlie the enhancement of cognitive abilities in brain evolution. The evolution of human cognition is then expected to have accompanied a prolongation of net neural-processing time due to the accumulation of processing time of individual neurons over an expanded number of neurons. Here, we confirmed this prediction and quantified the amount of prolongation in vivo, using noninvasive measurements of brain responses to sounds in unanesthetized human and nonhuman primates. Latencies of the N1 component of auditory-evoked potentials recorded from the scalp were approximately 40, 50, 60, and 100 ms for the common marmoset, rhesus monkey, chimpanzee, and human, respectively. Importantly, the prominent increase in human N1 latency could not be explained by the physical lengthening of the auditory pathway, and therefore reflected an extended dwell time for auditory cortical processing. A longer time window for auditory cortical processing is advantageous for analyzing time-varying acoustic stimuli, such as those important for speech perception. A novel hypothesis concerning human brain evolution then emerges: the increase in cortical neuronal number widened the timescale of sensory cortical processing, the benefits of which outweighed the disadvantage of slow cognition and reaction

Pathologic stage I non–small cell lung cancer with high levels of preoperative serum carcinoembryonic antigen: Clinicopathologic characteristics and prognosis

ObjectiveSurgery alone remains the standard therapy for patients with stage I non–small cell lung cancer. Although the preoperative serum level of carcinoembryonic antigen has been shown to be an independent prognostic factor, it has not yet been included in the staging system and does not alter the treatment strategy, especially in the selection of patients for adjuvant chemotherapy.MethodsFrom 1986 to 2003, preoperative and postoperative serum carcinoembryonic antigen levels were measured in 455 patients with completely resected pathologic stage I non–small cell lung cancer. We compared the clinicopathologic characteristics and outcomes among patients who had preoperative serum carcinoembryonic antigen levels within the normal range (N group, n = 323), patients who had high carcinoembryonic antigen levels before surgery but normal levels after surgery (HN group, n = 112), and patients who had high carcinoembryonic antigen levels before and after surgery (HH group, n = 20).ResultsThe significant characteristics of the HN group included the male sex, greater age, smoking, squamous cell histology, T2 status, lymphatic invasion, vascular invasion, and pleural invasion. Adenocarcinomas in patients of the HN group were more likely to be moderately to poorly differentiated. The 5-year survivals in the HN and HH groups were significantly lower (56.2% and 43.1%, respectively) than those in the N group (85.9%). Multivariate analysis revealed that greater age, non-adenocarcinoma histology, pleural invasion, and the carcinoembryonic antigen in the HN and HH groups were independent prognostic factors.ConclusionPatients with resected pathologic stage I non–small cell lung cancer and high preoperative serum carcinoembryonic antigen levels are a subgroup with a distinctly poor prognosis who display smoking-related clinicopathologic characteristics

Clinical Evaluation of the Peroral Cholangioscopy Using a New Videoscope

Peroral cholangioscopy (PCS) has been performed in 22 cases using XCHF-B200 (Olympus Optical Co.) since June 1995 and in 77 cases using CHF-B20 (Olympus Optical Co.) after EST from Jan. 1989. XCHF-B200 has a longer rigid portion of distal end and a smaller channel diameter than CHF-B20. The successful rate of PCS using XCHF-B200 (82%) was lower than that of CHF-B20 (89%). The vascular pattern and fine vertical groove of the bile duct mucosa were shown more clearly on the photographs obtained with XCHF-B200 than those obtained with CHF-B20. However, not enough biopsy specimens could be obtained because the channel diameter of XCHF-B200 was rather small

An attempt of dissemination of potential fishing zones prediction map of Japanese common squid in the coastal water, southwestern Hokkaido, Japan

Accurate prediction of potential fishing zones is regarded as one of the most immediate and effective approaches in operational fisheries. It helps fishermen reduce their cost on fuel and also decrease the uncertainty of their fish catches. To predict potential fishing zones of Japanese common squid, we derived fishing positions from the Defense Meteorological Satellite Program (DMSP) Operational Linescan System (OLS), combine with bathymetry and model-derived environmental factors from the 4D-VAR data assimilation system and fitted using habitat suitability index (HSI) model. Validations with an independent DMSP/OLS dataset showed better performance of the model in figuring out the squid aggregations than our previous model established with satellite-derived environmental data. Nighttime visible images during June and early July of 2013 derived from Day/Night band (DNB) of Visible Infrared Imaging Radiometer Suite (VIIRS) sensor with a better resolution and quality compared to DMSP/OLS, were also applied for validation and results showed differences of fitness between actual fishing activities and predictions in Japan Sea and Tsugaru Strait

A phase 3 randomized controlled trial of a COVID-19 recombinant vaccine S-268019-b versus ChAdOx1 nCoV-19 in Japanese adults

We assessed S-268019-b, a recombinant spike protein vaccine with a squalene-based adjuvant, for superiority in its immunogenicity over ChAdOx1 nCoV-19 vaccine among adults in Japan. In this multicenter, randomized, observer-blinded, phase 3 study, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)–naïve participants (aged ≥ 18 years, without prior infection or vaccination against SARS-CoV-2) were randomized (1:1) to receive either S-268019-b or ChAdOx1 nCoV-19 as two intramuscular injections given 28 days apart. Participants who provided consent for a booster administration received S-268019-b at Day 211. The primary endpoint was SARS-CoV-2 neutralizing antibody (NAb) titer on Day 57; the key secondary endpoint was the seroconversion rate for SARS-CoV-2 NAb titer on Day 57. Other endpoints included anti–SARS-CoV-2 S-protein immunoglobulin (Ig)G antibody titer and safety. The demographic and baseline characteristics were generally comparable between S-268019-b (n = 611) and ChAdOx1 nCoV-19 (n = 610) groups. S-268019-b showed superior immunogenicity over ChAdOx1 nCoV-19, based on their geometric mean titers (GMTs) and GMT ratios of SARS-CoV-2 NAb on Day 57 by cytopathic effect assay (GMT [95% confidence interval {CI}] 19.92 [18.68, 21.23] versus 3.63 [3.41, 3.87]; GMT ratio [95% CI] 5.48 [5.01, 6.00], respectively; two-sided p-values < 0.0001). Additionally, NAb measured using a cell viability assay also showed similar results (GMT [95% CI] 183.25 [168.04, 199.84] versus 24.79 [22.77, 27.00]; GMT ratio [95% CI] 7.39 [6.55, 8.35] for S-268019-b versus ChAdOx1 nCoV-19, respectively; p < 0.0001). The GMT of anti–SARS-CoV-2 S-protein IgG antibody was 370.05 for S-268019-b versus 77.92 for ChAdOx1 nCoV-19 on Day 57 (GMT ratio [95% CI] 4.75 [4.34, 5.20]). Notably, immune responses were durable through the end of the study. S-268019-b elicited T-helper 1 skewed T-cell response, comparable to that of ChAdOx1 nCoV-19. After the first dose, the incidence of solicited systemic treatment-related adverse events (TRAEs) was higher in the ChAdOx1 nCoV-19 group, but after the second dose, the incidence was higher in the S-268019-b group. Headache, fatigue, and myalgia were the most commonly reported solicited systemic TRAEs, while pain at the injection site was the most frequently reported solicited local TRAE following both doses in both groups. No serious treatment-related adverse serious TRAEs events were reported in the two groups. S-268019-b was more immunogenic than ChAdOx1 nCoV-19 vaccine and was well tolerated (jRCT2051210151)