Diabetes and its associated complications have increasingly become major challenges for global healthcare. The current therapeutic strategies involve insulin replacement therapy for type 1 diabetes (T1D) and small-molecule drugs for type 2 diabetes (T2D). Despite these advances, the complex nature of diabetes necessitates innovative clinical interventions for effective treatment and complication prevention. Accumulative evidence suggests that protein post-translational modifications (PTMs), including glycosylation, phosphorylation, acetylation, and SUMOylation, play important roles in diabetes and its pathological consequences. Therefore, the investigation of these PTMs not only sheds important light on the mechanistic regulation of diabetes but also opens new avenues for targeted therapies. Here, we offer a comprehensive overview of the role of several PTMs in diabetes, focusing on the most recent advances in understanding their functions and regulatory mechanisms. Additionally, we summarize the pharmacological interventions targeting PTMs that have advanced into clinical trials for the treatment of diabetes. Current challenges and future perspectives are also provided.

Visceral leishmaniasis (VL) or kala-azar is a vector-borne dreaded protozoal infection that is caused by the parasite Leishmania donovani. With increases in the dramatic infection rates, present drug toxicity, resistance, and the absence of an approved vaccine, the development of new antileishmanial compounds from plant sources remains the keystone for the control of visceral leishmaniasis. In this study, we evaluated the leishmanicidal effect of thymoquinone against L. donovani with an in vitro and ex vivo model. Thymoquinone exhibited potent antipromastigote activity with IC₅₀ and IC₉₀ concentrations achieved at 6.33 ± 1.21 and 20.71 ± 2.15 μM, respectively, whereas the IC₅₀ and IC₉₀ concentrations were found to be 7.83 ± 1.65 and 27.25 ± 2.20 μM against the intramacrophagic form of amastigotes, respectively. Morphological changes in promastigotes and growth reversibility study following treatment confirmed the leishmanicidal effect of thymoquinone. Further, thymoquinone exhibited leishmanicidal activities against L. donovani promastigote through cytoplasmic shrinkage, membrane blebbing, chromatin condensation, cellular and nuclear shrinkage, and DNA fragmentation, as observed under scanning and transmission electron microscopy analyses. The antileishmanial activity was exerted via programmed cell death as proved by exposure of phosphatidylserine, DNA nicking by TUNEL assay, and loss of mitochondrial membrane potential. Thymoquinone at a concentration of 200 μM was devoid of any cytotoxic effects against mammalian macrophage cells. Thymoquinone showed strong leishmanicidal activity against L. donovani, which is mediated via an apoptosis mode of parasitic cell death, and accordingly, thymoquinone may be the source of a new lead molecule for the cure of VL.

The re-emergence of Chikungunya virus (CHIKV) infection in humans with no approved antiviral therapies or vaccines is one of the major problems with global significance. In the present investigation, we screened 80 in-house quinoline derivatives for their anti-CHIKV activity by computational techniques and found 4-hydroxy-1-methyl-3-(3-morpholinopropanoyl)quinoline-2(1H)-one (QVIR) to have potential binding affinities with CHIKV nsP2 and E2 glycoproteins. QVIR was evaluated in vitro for its anti-CHIKV potential. QVIR showed strong inhibition of CHIKV infection with an EC₅₀ (50% effective concentration) value of 2.2 ± 0.49 μM without significant cytotoxicity (CC₅₀ > 200 μM) and was chosen for further elucidation of its antiviral mechanism. The infectious viral particle formation was abolished by approximately 72% at a QVIR concentration of 20 μM during infection in the BHK-21 cell line, and the CHIKV RNA synthesis was diminished by 84% for nsP2 as well as 74% for E2, whereas the levels of viral proteins were decreased by 69.9% for nsP2 and 53.9% for E2. Flow cytometry analysis confirmed a huge decline in the expression of viral nsP2 and E2 proteins by 71.84 and 67.7%, respectively. Time of addition experiments indicated that QVIR inhibited viral infection at early and late stages of viral replication cycle, and the optimal inhibition was observed at 16 h post infection. The present study advocates for the first time that QVIR acts as a substantial and potent inhibitor against CHIKV and might be as an auspicious novel drug candidate for the development of therapeutic agents against CHIKV infections.

The global burden of disease caused by a respiratory syncytial virus (RSV) is becoming more widely recognized in young children and adults. Heparan sulfate helps in attaching the virion through G protein with the host cell membrane. In this study, we examined the structural changes of ectodomain G protein (edG) in a wide pH range. The absorbance results revealed that protein maintains its tertiary structure at physiological and highly acidic and alkaline pH. However, visible aggregation of protein was observed in mild acidic pH. The intrinsic fluorescence study shows no significant change in the λ_max except at pH 12.0. The ANS fluorescence of edG at pH 2.0 and 3.0 forms an acid-induced molten globule-like state. The denaturation transition curve monitored by fluorescence spectroscopy revealed that urea and GdmCl induced denaturation native (N) ↔ denatured (D) state follows a two-state process. The fluorescence quenching, molecular docking, and 50 ns simulation measurements suggested that heparan sulfate showed excellent binding affinity to edG. Our binding study provides a preliminary insight into the interaction of edG to the host cell membrane via heparan sulfate. This binding can be inhibited using experimental approaches at the molecular level leading to the prevention of effective host–pathogen interaction.

Respiratory syncytial virus (RSV) is a leading viral pathogen causing acute lower respiratory tract infection in children. The G protein of RSV is involved in attachment with the host cell. It is a neutralizing antigen and thus a vaccine candidate. Heparan sulfate is a type of glycosaminoglycan (GAG) present on the host cell membrane that is involved in attachment with the G protein of RSV. We describe a novel approach for efficient expression and purification of the ectodomain G protein in the prokaryotic system and its biophysical characterization. The native ectodomain G protein was purified using a two-step process by Ni-NTA and DEAE weak anion-exchange chromatography through the supernatant obtained after cell lysis. In addition, the denatured form of the protein was also purified from the solubilized inclusion bodies (IBs) by Ni-NTA affinity chromatography with a higher yield. Dynamic light scattering (DLS) was performed to confirm the homogeneity of the purified protein. The effect of pH on the stability and structure of the purified protein was studied by circular dichroism (CD), fluorescence, and absorbance spectroscopy techniques. Isothermal titration calorimetry (ITC) and microscale thermophoresis (MST) were exploited to demonstrate the interaction of heparan sulfate with the ectodomain G protein. The dynamic light scattering results showed that the purified protein was homogenic and had a well-folded native conformation. Biophysical characterization of the protein revealed that it was stable and had intact secondary and tertiary structures at pH 7.5. CD analysis revealed that the protein showed a loss in the secondary structure at pH values 5.5 and 3.5, while absorbance spectroscopy suggested a stable tertiary structure at pH values 7.5 and 5.5 with a probable aggregation pattern at pH 3.5. This loss in the structure of the ectodomain G protein at low pH can be correlated with its physiological activity. A slight change in pH might play a crucial role in host–pathogen interactions. The fluorescence intensity of the protein decreased on moving toward a lower pH with no spectral shift in emission maxima. In addition, isothermal titration calorimetry and microscale thermophoresis results showed strong binding affinity of the ectodomain G protein with heparan sulfate. The binding of heparan sulfate with protein was probably due to the electrostatic interaction of positively charged amino acid residues of the heparin-binding domain of the protein and the negatively charged group of GAGs. Future studies may involve the development of possible therapeutic agents interacting with the G protein and affecting the overall charge and pH that might hinder the host–pathogen interaction.