The in vivo assembly and function of the N- and C-terminal halves of the Tn 10-encoded TetA protein in Escherichia coli

AbstractThe tetA gene was cut into its N- and C-terminal halves at the central EcoRI site and the two halves were subcloned individually or together under a separate lac promoter/operator. The expression of the C-terminal half was detected with a C-terminal-specific antibody. The amount of the N-terminal half in the cytoplasmic membrane was not affected by the presence of the C-terminal half. In contrast, the amount of the C-terminal half in the membrane was increased in the presence of the N-terminal half, indicating that the N-terminal half helps the stable folding of the C-terminal half in the membrane. Each half individually showed no tetracycline transport activity, however, when both halves were expressed together, the resultant complex showed about 40% of the tetracycline transport activity of the wild-type per number of the C-terminals of TetA protein in the membrane

The effect of lipopolysaccharide on lipid bilayer permeability of β-lactam antibiotics

AbstractThe lipid-bilayer permeability of cephalosporins was extensively suppressed by addition of lipopolysaccharide to liposomal membrane in proportion to the hydrophobicity of the drugs. This suggests that the polysaccharide chain layer contributes to the barrier function. The importance of the polysaccharide chain in the barrier function was also supported by the fact that the permeability to Rd-type lipopolysaccharide-containing liposomes showed essentially the same dependency on the hydrophobicity of the cephalosporins as that of the lipopolysaccharide-free liposomes. In this case the permeability of the cephalosporins was proportional to their hydrophobicity. Similar lipopolysaccharide effect was also observed in the permeation of penicillins

Substrate-induced acceleration of N-ethylmaleimide reaction with the Cys-65 mutant of the transposon Tn 10-encoded metal-tetracycline/H+ antiporter depends on the interaction of Asp-66 with the substrate

AbstractWe previously reported that the reaction of [14C]N-ethylmaleimide (NEM) with the S65C mutant of the transposon Tn10-encoded metal-tetracycline/H+ antiporter (TetA(B)) is competitively inhibited by tetracycline [Yamaguchi, A. et al., FEBS Lett. 322 (1993) 201–204]. However, this observation has been revealed to be a mistake. The reaction of [14C]NEM with S65C TetA(B) was significantly and reproducibly accelerated by tetracycline, i.e. not inhibited. When Asp-66 was replaced by Ala, the reaction of NEM with the Cys-65 residue was no longer affected by tetracycline. In contrast, when Arg-70 was replaced by Ala, the acceleration of the reaction was unaltered. The tetracycline acceleration of the reaction to the Cys-65 residue was further stimulated with energization of the membrane on the addition of NADH. On the other hand, the tetracycline-induced acceleration was not observed in the absence of a divalent cation. These observations indicated that the Cys-65 locus is exposed to the medium according to the interaction of a divalent cationtetracycline chelation complex with Asp-66

A novel compound, 1,1-dimethyl-5-(1-hydroxypropyl)-4,6,7-trimethylindan, is an effective inhibitor of the tet(K) gene-encoded metal-tetracycline/H+ antiporter of Staphylococcus aureus

AbstractA novel indan derivative, 1,1-dimethyl-5-(1-hydroxypropyl)-4,6,7-trimethylindan (Ro 07-3149), was found to be a strong inhibitor of the tet(K) gene-encoded tetracycline/H+ antiporter of Staphylococcus aureus. One micromole of this compound per mg membrane protein was enough for complete inhibition of the Tet(K)-mediated tetracycline transport and tetracycline-coupled proton transport, without the energy state of the membrane being affected. The mode of inhibition was non-competitive. Although this compound caused membrane de-energization at a high concentration, the IC50 value for de-energization (7.3 μmol/mg membrane protein) was about 17 times and 33 times higher than the values for Tet(K)-mediated proton/tetracycline antiport and [3H]tetracycline transport, respectively, indicating that the inhibitory action of Ro 07-3149 is not due to the uncoupling effect of the inhibitor

Oncofertility care in young women and the outcomes of pregnancy over the last 5 years

Aim: To ascertain the actual outcomes of oncofertility care in young women to provide more appropriate care. Materials & methods: We analyzed the data of 67 female patients under 43 years of age who underwent oncofertility care between January 2015 and September 2019. Results: There were 28 patients with breast cancer, 19 patients with hematologic cancer and 20 patients with other cancer diagnoses. Breast cancer patients tended to take longer than hematologic cancer patients to initiate oncofertility treatment. Despite undergoing oncofertility care, seven of nine pregnant patients did not choose assisted reproductive technology (ART). Conclusion: As spontaneous pregnancies were more common than ART pregnancies in our study, pregnancy by not only ART but also non-ART method is a viable option for young cancer survivors

Development of healthy lifestyle consciousness index for gynecological cancer patients

PURPOSE: Healthy lifestyle is related to quality of life (QOL) after cancer diagnosis and prognosis. However, there are few reports on patients conscious of healthy lifestyle and patients requiring medical providers' attention regarding healthy lifestyle. We aimed to develop a healthy lifestyle consciousness index (HLCI) for cancer patients and evaluated its validity in gynecological cancer patients. METHODS: The HLCI was designed to assess degree of healthy lifestyle consciousness, including items regarding "diet, " "exercise, " "body weight, " and "sleep." Exploratory factor analysis was performed for dimensionality of the scale; Cronbach's alpha was calculated to assess internal-consistency reliability. For criterion-based validity, we calculated proportions of stage III/IV gynecological malignancies in those with categorized HLCI scores based on tertiles. Concurrent validity was evaluated between HLCI and other quality of life (QOL) scales including European Organization for Research and Treatment of Cancer QLQ-C30 in limited patients. RESULTS: HLCI comprised five 10-point items (0-45); higher values implied improved healthy lifestyle consciousness. Data from 108 gynecological malignancy patients at Kyoto University Hospital were analyzed. The mean age of subjects was 55.8 years; 36.1% of them had uterine corpus cancer; 34.3% were at stage III/IV of gynecological malignancy. The factor analysis revealed HLCI was unidimensional; the reliability based on Cronbach's alpha was satisfactory (0.88). The proportions of stage III/IV gynecological malignancies were 25.7%, 33.3%, and 44.4% in those with first (7-24 points), second (25-30 points), and third (31-46 points) tertiles of HLCI score, respectively. For patients with other QOL scales (n = 25), the mean scores of global health status of QLQ-C30 were 33.3, 50.0, and 83.3 for first, second, and third tertiles of HLCI score, respectively. CONCLUSION: HLCI was successfully validated; thus, patients with advanced stages or higher QOL might have strong consciousness regarding healthy lifestyle. HLCI may be useful in precision care for improved lifestyles and QOL

Dynamic action of an intrinsically disordered protein in DNA compaction that induces mycobacterial dormancy

抗酸菌における決定的な休眠誘導機構を発見 --天然変性タンパク質による新規のDNA凝集メカニズム--. 京都大学プレスリリース. 2024-02-14.Mycobacteria are the major human pathogens with the capacity to become dormant persisters. Mycobacterial DNA-binding protein 1 (MDP1), an abundant histone-like protein in dormant mycobacteria, induces dormancy phenotypes, e.g. chromosome compaction and growth suppression. For these functions, the polycationic intrinsically disordered region (IDR) is essential. However, the disordered property of IDR stands in the way of clarifying the molecular mechanism. Here we clarified the molecular and structural mechanism of DNA compaction by MDP1. Using high-speed atomic force microscopy, we observed that monomeric MDP1 bundles two adjacent DNA duplexes side-by-side via IDR. Combined with coarse-grained molecular dynamics simulation, we revealed the novel dynamic DNA cross-linking model of MDP1 in which a stretched IDR cross-links two DNA duplexes like double-sided tape. IDR is able to hijack HU function, resulting in the induction of strong mycobacterial growth arrest. This IDR-mediated reversible DNA cross-linking is a reasonable model for MDP1 suppression of the genomic function in the resuscitable non-replicating dormant mycobacteria

Low-Grade Endometrial Stromal Sarcoma with a Nodule-in-Nodule Appearance in Preoperative Magnetic Resonance Images

Low-grade endometrial stromal sarcoma (LG-ESS) is a rare malignant disease and demonstrates various patterns in preoperative imaging. Therefore, accurate diagnosis is important. Given its unique form, we report a case of LG-ESS with a nodule-in-nodule appearance on preoperative imaging. A 41-year-old woman was referred to our department for further examination of a 45 mm diameter uterine corpus mass. Preoperative magnetic resonance imaging (MRI) revealed several small nodules within a larger nodule. T2-weighted images showed moderate-to-high signal intensity with focal bands of low signal intensity in the small nodules. The patient underwent total abdominal hysterectomy and bilateral salpingo-oophorectomy. Histopathological findings of the small nodules showed densely concentrated endometrial stromal cells reminiscent of a proliferative phase endometrium with a concentric arrangement of small spiral arteriole-like vessels. The small nodules exhibited an expansile growth pattern and were surrounded by less densely concentrated endometrial stromal cells intermingled with the normal uterine myometrium. LG-ESS with smooth muscle differentiation and sex cord-like elements was partially observed. In summary, LG-ESS demonstrating a unique nodule-in-nodule appearance on preoperative imaging histopathologically comprised tumor cells of varying densities. Our current case suggests that preoperative diagnostic imaging with MRI may be useful