Targeting dermatophyte Cdc42 and Rac GTPase signaling to hinder hyphal elongation and virulence.

The development of antifungal drugs requires novel molecular targets due to limited treatment options and drug resistance. Through chemical screening and establishment of a novel genetic technique to repress gene expression in Trichophyton rubrum, the primary causal fungus of dermatophytosis, we demonstrated that fungal Cdc42 and Rac GTPases are promising antifungal drug targets. Chemical inhibitors of these GTPases impair hyphal formation, which is crucial for growth and virulence in T. rubrum. Conditional repression of Cdc24, a guanine nucleotide exchange factor for Cdc42 and Rac, led to hyphal growth defects, abnormal cell morphology, and cell death. EHop-016 inhibited the promotion of the guanine nucleotide exchange reaction in Cdc42 and Rac by Cdc24 as well as germination and growth on the nail fragments of T. rubrum and improved animal survival in an invertebrate infection model of T. rubrum. Our results provide a novel antifungal therapeutic target and a potential lead compound.

The Ptk2-Pma1 pathway enhances tolerance to terbinafine in Trichophyton rubrum.

The increasing prevalence of dermatophyte resistance to terbinafine, a key drug in the treatment of dermatophytosis, represents a significant obstacle to treatment. Trichophyton rubrum is the most commonly isolated fungus in dermatophytosis. In T. rubrum, we identified TERG_07844, a gene encoding a previously uncharacterized putative protein kinase, as an ortholog of budding yeast Saccharomyces cerevisiae polyamine transport kinase 2 (Ptk2), and found that T. rubrum Ptk2 (TrPtk2) is involved in terbinafine tolerance. In both T. rubrum and S. cerevisiae, Ptk2 knockout strains were more sensitive to terbinafine compared with the wild types, suggesting that promotion of terbinafine tolerance is a conserved function of fungal Ptk2. Pma1 is activated through phosphorylation by Ptk2 in S. cerevisiae. Overexpression of T. rubrum Pma1 (TrPma1) in T. rubrum Ptk2 knockout strain (ΔTrPtk2) suppressed terbinafine sensitivity, suggesting that the induction of terbinafine tolerance by TrPtk2 is mediated by TrPma1. Furthermore, omeprazole, an inhibitor of plasma membrane proton pump Pma1, increased the terbinafine sensitivity of clinically isolated terbinafine-resistant strains. These findings suggest that, in dermatophytes, the TrPtk2-TrPma1 pathway plays a key role in promoting intrinsic terbinafine tolerance and may serve as a potential target for combinational antifungal therapy against terbinafine-resistant dermatophytes.

Serum MMP3 and IL1-RA levels may be useful biomarkers for detecting asthma and chronic obstructive pulmonary disease overlap in patients with asthma.

Background: Asthma and chronic obstructive pulmonary disease (COPD) overlap (ACO) is characterized by concurrent features of asthma and COPD. Since disease pathogenesis, severities, and treatments differ between asthma and ACO, it is important to differentiate them. Objective: To clarify and compare the characteristics of ACO and asthma and identify the serum biomarkers for differentiating them, especially in older patients. Methods: This study used the data of 639 participants from the nationwide cohort study, the NHOM-Asthma study, an asthma registry in Japan, with complete information on smoking history, respiratory function, and serum biomarkers. ACO was defined as the self-reported comorbidity of COPD or emphysema, or with obstructive pulmonary function and smoking history (pack-years≥10). The clinical characteristics of patients with ACO and asthma without COPD were compared. The serum biomarkers for differentiation were examined using receiver operating characteristic curves and multivariable analysis. The associations between the biomarkers and age were also analyzed. Results: Of the 639 asthma patients, 125 (19.6%) were diagnosed with ACO; these patients were older and male-dominant and had a higher prevalence of comorbidities such as hypertension, diabetes, and stroke. Among the serum biomarkers that were significantly different between ACO and asthma without COPD, the YKL-40/CHI3L1, MMP3, and IL-1RA levels showed a high area under the curve for discriminating ACO. Only the MMP3 and IL-1RA levels were significantly higher among ACO patients, regardless of age and sex; the YKL-40/CHI3L1 levels were not different due to the effect of age. Conclusion: MMP3 and IL-1RA may be useful serum biomarkers for distinguishing ACO from asthma.

TrCla4 promotes actin polymerization at the hyphal tip and mycelial growth in Trichophyton rubrum.

IMPORTANCE: Superficial fungal infections, such as athlete's foot, affect more than 10% of the world's population and have a significant impact on quality of life. Despite the fact that treatment-resistant fungi are a concern, there are just a few antifungal drug targets accessible, as opposed to the wide range of therapeutic targets found in bacterial infections. As a result, additional alternatives are sought. In this study, we generated a PAK TrCla4 deletion strain (∆Trcla4) of Trichophyton rubrum. The ∆Trcla4 strain exhibited deficiencies in mycelial growth, hyphal morphology, and polarized actin localization at the hyphal tip. IPA-3 and FRAX486, small chemical inhibitors of mammalian PAK, were discovered to limit fungal mycelial proliferation. According to our findings, fungal PAKs are interesting therapeutic targets for the development of new antifungal medicines.

Doping of molecular semiconductors through proton-coupled electron transfer.

The chemical doping of molecular semiconductors is based on electron-transfer reactions between the semiconductor and dopant molecules; here, the redox potential of the dopant is key to control the Fermi level of the semiconductor1,2. The tunability and reproducibility of chemical doping are limited by the availability of dopant materials and the effects of impurities such as water. Here we focused on proton-coupled electron-transfer (PCET) reactions, which are widely used in biochemical processes3,4; their redox potentials depend on an easily handled parameter, that is, proton activity. We immersed p-type organic semiconductor thin films in aqueous solutions with PCET-based redox pairs and hydrophobic molecular ions. Synergistic reactions of PCET and ion intercalation resulted in efficient chemical doping of crystalline organic semiconductor thin films under ambient conditions. In accordance with the Nernst equation, the Fermi levels of the semiconductors were controlled reproducibly with a high degree of precision-a thermal energy of about 25 millielectronvolts at room temperature and over a few hundred millielectronvolts around the band edge. A reference-electrode-free, resistive pH sensor based on this method is also proposed. A connection between semiconductor doping and proton activity, a widely used parameter in chemical and biochemical processes, may help create a platform for ambient semiconductor processes and biomolecular electronics.

Serum IgA augments adhesiveness of cultured lung microvascular endothelial cells and suppresses angiogenesis.

Immunoglobulin A (IgA) is important in local immunity and is also abundant in the blood. This study aimed to evaluate the effects of serum IgA on cultured lung microvascular endothelial cells (HMVEC-Ls), which are involved in the pathogenesis of inflammatory lung diseases. Serum IgA induced adhesion molecules and inflammatory cytokine production from HMVEC-Ls, and enhanced adhesion of peripheral blood mononuclear cells to HMVEC-Ls. In contrast, migration, proliferation, and tube formation of HMVEC-Ls were significantly suppressed by serum IgA. Experiments with siRNAs and western blotting revealed that two known IgA receptors, ß1,4-galactosyltransferase 1 (b4GALT1) and asialoglycoprotein receptor 1 (ASGR1), and mitogen-activated protein kinase and nuclear factor-kappa B pathways were partly involved in serum IgA-induced cytokine production by HMVEC-Ls. Collectively, serum IgA enhanced cytokine production and adhesiveness of HMVEC-L, with b4GALT1 and ASGR1 partially being involved, and suppressed angiogenesis. Thus, serum IgA may be targeted to treat inflammatory lung diseases.

Mechanical Tuning of Aggregated States for Conformation Control of Cyclized Binaphthyl at the Air-Water Interface.

An air-water interface enables molecular assemblies and conformations to be controlled according to their intrinsic interactions and anisotropic stimuli. The chirality and conformation of binaphthyl derivatives have been controlled by tuning molecular aggregated states in solution. In this study, we have tuned molecular aggregated states of monobinaphthyldurene (MBD) by applying different mechanical stimuli to control the conformation at the air-water interface. Density functional theory calculations indicate that MBD exists essentially in two conformations, namely, 1-MBD (most stable) and 2-MBD (less stable). MBD was mechanically dissolved in appropriate lipid matrices using the Langmuir-Blodgett (LB) method, while pure MBD was self-assembled at the dynamic air-water interface in the absence of or by applying vortex motions (vortex LB method). In MBD mixed monolayer, surface pressure-molecular area measurements and atomic force microscopy observations suggest that separate lipids and MBD phases transform to mixed phases induced by the dissolution of MBD into the lipid matrices during mechanical compression at the air-water interface. Circular dichroism measurements indicate that molecular conformation changes from 1-MBD to 2-MBD in passing from a separated phase to a mixed MBD/lipid phase. In addition, the molecular aggregated states and conformations of MBD depend on the spreading volume and vortex flow rate when applying the vortex LB method. Molecular conformations and aggregated states of MBD could be controlled continuously by applying a mechanical stimulus at the air-water interface.

epi-Aszonalenin B from Aspergillus novofumigatus inhibits NF-κB activity induced by ZFTA-RELA fusion protein that drives ependymoma.

Nuclear factor-kappa B (NF-κB) signaling is an intracellular signaling pathway involved in inflammatory responses and the pathogenesis of various cancers, including ependymoma, which is a rare and chemotherapy-resistant glioma. Several isoforms of fusion proteins that consist of a nuclear protein, zinc finger translocation associated (ZFTA), and RELA (ZFTA-RELA), an NF-κB-signaling effector transcription factor, cause excessive activation of the NF-κB signaling pathway and result in supratentorial ependymomas (ST-EPN-RELA). As inhibitors of NF-κB activity induced by ZFTA-RELA are expected to be therapeutic agents for ST-EPN-RELA, we established an NF-κB responsive luciferase reporter cell line that expresses the most common isoform of ZFTA-RELA in a doxycycline-dependent manner. Using this reporter cell line, we screened fungus extracts for compounds that inhibit the NF-κB activity induced by ZFTA-RELA expression and identified aszonalenin, an alkaloid from Aspergillus novofumigatus. We also purified analogs of aszonalenin, namely acetylaszonalenin and epi-aszonalenin B and C. In a luciferase assay using cells constitutively expressing luciferase (counter assay), acetylaszonalenin and epi-aszonalenin C showed non-specific inhibition of the luciferase activity. Aszonalenin and epi-aszonalenin B inhibited the NF-κB responsive luciferase activity by expressing ZFTA-RELA more strongly than the luciferase activity in the counter assay. The upregulation of endogenous NF-κB responsive genes, such as CCND1, ICAM1, and L1CAM, by ZFTA-RELA expression was inhibited by epi-aszonalenin B, but not by aszonalenin. This study suggests that epi-aszonalenin B may be a lead compound for the therapeutic development of ST-EPN-RELA.

Influence of oral health on frailty in patients with type 2 diabetes aged 75 years or older.

BACKGROUND: Poor oral health conditions are known to affect frailty in the older adults. Diabetes is a risk factor for both poor oral health and frailty, therefore, oral health status may affect frailty in diabetic patients more than in the general population. The purpose of this study was to evaluate the influence of oral health and other factors on frailty and the relationship among oral health, diabetes and frailty in older adult patients with type 2 diabetes. METHODS: Patients with type 2 diabetes aged 75 years or older were included in this cross-sectional study. Eligible patients were surveyed by questionnaire for frailty, oral health status, and cognitive and living functions. Factors influencing pre-frailty, frailty, and individual frailty screening index (FSI) classes were evaluated. RESULTS: Of the 111 patients analyzed, 66 cases (59.5%) were categorized as robust, 33 cases (29.7%) as pre-frailty, and 12 cases (10.8%) as frailty. The oral frailty index, the cognitive and living functions score, and BMI were found to be factors influencing pre-frailty or frailty. In the evaluation of individual FSI classes, BMI had an influence on those with a FSI ≤2. The cognitive and living functions score was a factor influencing those with FSI ≤3. The oral frailty index was found to have a significant influence on all FSI classes. CONCLUSIONS: Poor oral health has an influence on frailty in patients with type 2 diabetes aged ≥75. In this patient population, as frailty progresses, the impact of oral health on frailty may increase. TRIAL REGISTRATION: This study was retrospectively registered in UMIN-CTR ( UMIN000044227 ).

Ependymoma associated protein Zfta is expressed in immature ependymal cells but is not essential for ependymal development in mice.

The fusion protein of uncharacterised zinc finger translocation associated (ZFTA) and effector transcription factor of tumorigenic NF-κB signalling, RELA (ZFTA-RELA), is expressed in more than two-thirds of supratentorial ependymoma (ST-EPN-RELA), but ZFTA's expression profile and functional analysis in multiciliated ependymal (E1) cells have not been examined. Here, we showed the mRNA expression of mouse Zfta peaks on embryonic day (E) 17.5 in the wholemount of the lateral walls of the lateral ventricle. Zfta was expressed in the nuclei of FoxJ1-positive immature E1 (pre-E1) cells in E18.5 mouse embryonic brain. Interestingly, the transcription factors promoting ciliogenesis (ciliary TFs) (e.g., multicilin) and ZFTA-RELA upregulated luciferase activity using a 5' upstream sequence of ZFTA in cultured cells. Zftatm1/tm1 knock-in mice did not show developmental defects or abnormal fertility. In the Zftatm1/tm1 E1 cells, morphology, gene expression, ciliary beating frequency and ependymal flow were unaffected. These results suggest that Zfta is expressed in pre-E1 cells, possibly under the control of ciliary TFs, but is not essential for ependymal development or flow. This study sheds light on the mechanism of the ZFTA-RELA expression in the pathogenesis of ST-EPN-RELA: Ciliary TFs initiate ZFTA-RELA expression in pre-E1 cells, and ZFTA-RELA enhances its own expression using positive feedback.

Breathing irregularities before sleep onset on polysomnography in patients with heart diseases.

PURPOSE: Severe cardiac dysfunction can manifest with diurnal breathing irregularity. However, it remains to be clarified whether or not diurnal breathing irregularity is observed in patients with heart diseases, including relatively mild chronic heart failure (CHF), compared to those without heart diseases. METHODS: In this cross-sectional study, consecutive inpatients who were admitted for evaluation of sleep-disordered breathing were enrolled. We extracted 3.5 min of stable respiratory signals before sleep onset using polysomnography, analyzed the airflow data using fast Fourier transform, and quantified breathing irregularities using Shannon entropy S. RESULTS: A total of 162 subjects were evaluated. Among these, 39 subjects had heart diseases, including ischemic heart disease (IHD), atrial fibrillation (Af), CHF, and a history of aortic dissection. The values of Shannon entropy S of airflow signals in subjects with heart diseases were significantly higher than in those without heart diseases (p < 0.001). After excluding CHF, the Shannon entropy S was also significantly higher in subjects with heart diseases than in those without heart diseases (p < 0.001). The values of Shannon entropy S were significantly correlated with plasma brain natriuretic peptide levels (r = 0.443, p < 0.001). Although the values were also significantly correlated with body mass index, the presence of heart diseases was independently associated with breathing irregularity in the multiple logistic analysis. Matching analysis revealed consistent differences between subjects with heart diseases and without heart diseases. CONCLUSION: Breathing irregularity was observed before sleep onset in subjects with heart diseases who underwent polysomnography to diagnose sleep-disordered breathing.

Utility of a shortened Hasegawa Dementia Scale Revised questionnaire to rapidly screen and diagnose Alzheimer's disease.

AIMS: The aim of this study was to analyze the sensitivity and specificity of a shortened Hasegawa Dementia Scale Revised (shortened HDS-R) questionnaire and explore its utility for the rapid screening and diagnosis of Alzheimer's disease (AD). METHODS: We included 113 patients over the age of 60 years who visited our hospital from June 2018 to January 2021 including 70 subjects with AD and 43 healthy subjects. AD was diagnosed in accordance with the diagnostic criteria of the Diagnostic and Statistical Manual of Mental Disorders, 4th edition, and the standard HDS-R questionnaire was used as a neuropsychological examination. The shortened HDS-R questionnaire was composed of the first seven subdomains (1 to 7) of the HDS-R questionnaire and excluded subdomains 8 and 9. Magnetic resonance imaging (MRI) was performed to calculate the degree of atrophy of the whole brain, hippocampus, and parahippocampal gyrus. RESULTS: The cumulative contribution ratio of subdomains 1 to 7 of the HDS-R questionnaire was as high as 94%, indicating that the construct validity of the shortened HDS-R was very good. The correlation coefficient of the total scores of the shortened HDS-R and the HDS-R was 0.96, indicating that the criterion-related validity was also very good. Furthermore, the shortened HDS-R was significantly negatively correlated with the degree of atrophy in the whole brain, hippocampus, and parahippocampal gyrus, indicating that its concurrent validity was very good in relation to imaging parameters. Cronbach's α coefficient of the shortened HDS-R was 0.76, and the correlation coefficient of the item-total correlation analysis was between 0.68 and 0.76, indicating that this questionnaire has high internal consistency and reliability. The total shortened HDS-R score of the normal group (17.0 ± 1.9) was significantly higher than that of the AD group (8.6 ± 3.8), demonstrating that the total shortened HDS-R score can be used to identify healthy individuals and patients with AD. When the cutoff score was 14 of 15, the sensitivity was 92.9% and the specificity was 88.4%. The diagnostic ability of the shortened HDS-R was 91.2%, which indicates that it is similar to the full HDS-R questionnaire as an AD screening tool. CONCLUSION: As a neuropsychological examination questionnaire for the screening and diagnosis of AD, the shortened HDS-R had very high validity and reliability. Its sensitivity, specificity, and diagnostic ability were similar to those of the gold standard HDS-R; therefore, it can be considered a concise and useful questionnaire for AD screening and diagnosis in the older population.

Helicity Manipulation of a Double-Paddled Binaphthyl in a Two-Dimensional Matrix Field at the Air-Water Interface.

Molecular behavior and functionality are affected by their prevailing immediate environment. Molecular machines function according to conformational variations and have been studied largely in solution states. In order to access more highly complex functional molecular machines, it is necessary to analyze and control them in various environments. We have designed and synthesized a bisbinaphthyldurene (BBD) molecule that has two binaphthyl groups connected through a central durene moiety, allowing for the formation of several conformers. In density functional theory (DFT) calculations, BBD has five major conformers, denoted anti-1/anti-2/syn-1/syn-2/flat. It has been demonstrated that BBD exhibits different conformations in solution (anti-1 and syn-1) than on a gold surface (syn dimer and flat). In this work, the ratio of BBD conformations has been controlled in mixed monolayers with several different lipids at an air-water interface in order to compare conformational activity under different conditions. The conformations of BBD in transferred films obtained by using Langmuir-Blodgett techniques were estimated from circular dichroism spectra and DFT calculations. It has been found that the conformation of BBD in the mixed monolayer depends on its aggregated state, which has been controlled here by the mechanical properties and miscibility. In mixed monolayers with "hard" lipids having less miscibility with BBD as well as in cast film, BBD is self-aggregated and mostly forms stable anti-1 and syn-1 conformations, while unstable anti-2 and syn-2 conformers dominated in the more dispersed states involving "soft" lipids, which show good miscibility with BBD. Conformational changes in BBD are due to the formation of different aggregated states in each mixed monolayer according to the miscibility. Overall, BBD molecular conformations (and the resulting spectra) could be tuned by controlling the environment whether in solution, on a solid substrate, or in an admixture with lipids at the air-water interface.

Cognitive function has a stronger correlation with perceived age than with chronological age.

AIM: The perceived age of older adults, as measured by their facial appearance, has been shown to be a robust biomarker of aging predictive of survival, telomere length and DNA methylation, and reportedly correlates with carotid atherosclerosis and bone status. This study aimed to determine whether metrics of dementia, including general cognition, vitality, depressive state and self-supportability, have stronger correlations with perceived age than with chronological age. METHODS: This study included 124 patients who were admitted to the Department of Geriatric Medicine, The University of Tokyo Hospital, on account of being suspected of cognitive decline. The Mini-Mental State Examination, Vitality Index, Geriatric Depression Scale-15, instrumental activities of daily living and Barthel Index were carried out. Five experienced geriatricians and five experienced clinical psychologists determined the perceived age of participants based on photographs. RESULTS: The average values of the 10 raters showed excellent reliability (intraclass correlation coefficient (3, 10) = 0.941). Steiger's test revealed that perceived age showed a significantly better correlation with the Mini-Mental State Examination (female) and Vitality Index (total, female) than did chronological age, but not with Geriatric Depression Scale-15, instrumental activities of daily living or the Barthel Index. CONCLUSIONS: Perceived age was shown to be a reliable biomarker for cognitive assessment. Geriatr Gerontol Int 2020; 20: 779-784.

2D Nanoarchitectonics: Soft Interfacial Media as Playgrounds for Microobjects, Molecular Machines, and Living Cells.

Soft and flexible two-dimensional (2D) systems, such as liquid interfaces, would have much more potentials in dynamic regulation on nano-macro connected functions. In this Minireview article, we focus especially on dynamic motional functions at liquid dynamic interfaces as 2D material systems. Several recent examples are selected to be explained for overviewing features and importance of dynamic soft interfaces in a wide range of action systems. The exemplified research systems are mainly classified into three categories: (i) control of microobjects with motional regulations; (ii) control of molecular machines with functions of target discrimination and optical outputs; (iii) control of living cells including molecular machine functions at cell membranes and cell/biomolecular behaviors at liquid interface. Sciences on soft 2D media with motional freedom and their nanoarchitectonics constructions will have increased importance in future technology in addition to popular rigid solid 2D materials.

Publisher Correction: Characteristics of factors for decreased lung function in elderly patients with type 2 diabetes.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Characteristics of factors for decreased lung function in elderly patients with type 2 diabetes.

Chronic obstructive pulmonary disease (COPD) often accompanies type 2 diabetes mellitus (T2DM). However, background factors affecting these diseases in the elderly remain unclear. Eligible patients with T2DM were divided into two age groups-non-elderly (<65 years) and elderly (≥65 years); COPD, ratio of forced expiratory volume in one second to forced expiratory volume (FEV1/FVC ratio), and percent predicted forced expiratory volume in one second (FEV1% predicted) were examined, and factors related to reduced respiratory function according to age group were evaluated. In total, 371 patients with T2DM were analysed. COPD was found in 9 patients (5.3%) in the non-elderly group and 45 (22.5%) in the elderly group. In the elderly, male sex, low body mass index (BMI), insulin therapy, and high C-peptide immunoreactivity levels were factors related to COPD. In the non-elderly, age, female sex, high BMI were factors related to decreased FEV1% predicted. Female sex was factor related to decreased FEV1% predicted in both age groups. Low BMI was a factor related to reduced respiratory function in elderly patients and high BMI was a factor related to reduced respiratory function in non-elderly patients. Thus, BMI needs to be managed according to the age and general condition of T2DM patients.

Additive effects of Kothala himbutu (Salacia reticulata) extract and a lactic acid bacterium (Enterococcus faecalis YM0831) for suppression of sucrose-induced hyperglycemia in an in vivo silkworm evaluation system.

Using a silkworm evaluation system, we previously evaluated various substances that suppress postprandial hyperglycemia. Enterococcus faecalis YM0831, a lactic acid bacterium that inhibits glucose uptake by the human intestinal Caco-2 cell line, exhibited hyperglycemia-suppressing effects in the silkworm system. In the present study, we found that Kothala himbutu (Salacia reticulata) extract, a traditional medicine containing α-glucosidase inhibitors, suppressed sucrose-induced hyperglycemia in the silkworm system. Moreover, combined oral administration of lactic acid bacteria YM0831 with Kothala himbutu extract had stronger suppressive effects on sucrose-induced hyperglycemia than single administration of either component. These findings suggest that the silkworm system provides a simple way to evaluate the effects of supplements on the suppression of blood glucose level induced by sucrose ingestion.

N-Terminal (1→3)-ß-d-Glucan Recognition Proteins from Insects Recognize the Difference in Ultra-Structures of (1→3)-ß-d-Glucan.

Recognition of (1→3)-ß-d-glucans (BGs) by invertebrate ß-1,3-d-glucan recognition protein (BGRP) plays a significant role in the activation of Toll pathway and prophenoloxidase systems in insect host defense against fungal invasion. To examine the structure diversity of BGRPs for the recognition of physiochemically different BGs, the binding specificity of BGRPs cloned from four different insects to structure different BGs was characterized using ELISA. Recombinant BGRPs expressed as Fc-fusion proteins of human IgG1 bound to the solid phase of BGs. Based on the binding specificities, the BGRPs were categorized into two groups with different ultrastructures and binding characters; one group specifically binds BGs with triple-helical conformation, while the other group recognizes BGs with disordered conformations like single-helical or partially opened triple helix. The BGRPs from the silkworm and the Indian meal moth bound to the BGs with a triple-helical structure, whereas BGRPs from the red flour beetle and yellow mealworm beetle showed no binding to triple-helical BGs, but bound to alkaline-treated BGs that have a partially opened triple-helical conformation. This evidence suggests that the insect BGRPs can distinguish between different conformations of BGs and are equipped for determining the diversity of BG structures.