Current Proteomics - Volume 19, Issue 1, 2022

Self-attachment of proteins leading to the formation of highly insoluble protein oligomers and aggregates has become an important focus of research owing to its diverse implications in pathophysiology and diseases. This has become a more frequent phenomenon in most neurological and neurodegenerative diseases as well as in dementia. In recent years such an event of protein aggregation has been linked to other disease conditions, disorders or adverse health conditions. Interestingly, aggregation of protein also plays a role in development, growth or metabolism. Most often, physiological proteins are initially bio-synthesised in native or nascent geometrical forms or conformations, but later they undergo specific folding patterns and thereby acquire a stable configuration that is biologically relevant and active. It is highly important that these proteins remain in their biologically active configuration in order to exert their functional properties. Any alteration or change to this structural configuration can be detrimental to their specific functions and may cause pathological consequences leading to the onset of diseases or disorders. Several factors such as the action of chaperones, binding partners, physiological metal ions, pH level, temperature, ionic strength, interfacial exposure (solid-liquid, liquid-liquid, gas-liquid), mutation and post-translational modification, chemical changes, interaction with small molecules such as lipids, hormones, etc. and solvent environment have been either identified or proposed as important factors in conferring the ultimate status of protein structure and configuration. Among many misfolding protein conformations, self-assembly or aggregation is the most significant. It leads to the formation of highly oligomeric self-aggregates that precipitate and interfere with many biochemical processes with serious pathological consequences. The most common implication of protein aggregation leading to the formation of deposits / plaques of various morphological types is the onset of neurological and neurodegenerative diseases that include Alzheimer’s, Parkinson’s, Huntington, ALS (Amyotrophic Lateral Sclerosis), CJD (Creutzfeldt Jakob Dementia), Prion diseases, Amyloidosis and other forms of dementia. However, increasing studies have revealed that protein aggregation may also be associated with other diseases such as cancer, type 2 diabetes, renal, corneal and cardiovascular diseases. Protein aggregation diseases are now considered as part of “Proteinopathy” which refers to conditions where proteins become structurally abnormal or fail to fold into stable normal configurations. In this review, we reflect on various aspects of protein self-aggregation, potential underlying causes, mechanism, role of secondary structures, pathological consequences and possible intervention strategies as reported in published literature.

Background: Triple-negative breast cancer (TNBC) is a subtype of invasive cancer in breast with the symptoms of unfavourable prognosis and limited targeted treatment options. Evidence of changes in the metabolic status of TNBC, characterised by increased glycolysis, mitochondrial oxidative phosphorylation, as well as production and utilization of tricarboxylic acid cycle intermediates. Objective: The objective of this study is to investigate the proteins altered in cryptotanshinone treated MDA-MB-231 cells and explore the key pathways and specific molecular markers involved in cryptotanshinone treatment. Methods: We use unlabeled quantitative proteomics to gain insight into the anticancer mechanism of cryptotanshinone on MDA-MB231 triple negative breast cancer cells. And flow cytometry was used to detect apoptosis and changes in cell mitochondrial membrane potential. Results: We show that inhibiting the expression of electron transport chain complex proteins, also inhibits mitochondrial oxidative phosphorylation. Additionally, down-regulation of the ribosime biogenesis pathway was found to inhibit cell metabolism. Conclusion: In summary, results show that cryptotanshinone can trigger rapid and irreversible apoptosis in MDA-MB-231 cells through effectively inhibiting cell metabolism.

Background: Serum uric acid (UA) is positively correlated with bone mineral density (BMD). However, the mechanism by which serum UA affects BMD remains unclear. Objective: The aim was carried out to search for the functional proteins related to serum UA and femoral neck BMD to better understand the pathophysiological mechanism of osteoporosis. Materials and Methods: In this study, patients in the UA group (hyperuricaemia combined with femoral neck fracture) and the control group (normal uricaemia combined with femoral neck fracture) were selected according to the inclusion criteria. Total protein was extracted from the femoral neck of each patient. Fluorescence differential gel electrophoresis was used to separate the total proteins, and the differentially expressed protein spots were detected by image analysis. After enzyme digestion, peptide mass fingerprinting and database searches were performed to identify the differentially expressed proteins. DAVID software and Kyoto Encyclopedia of Genes and Genomes (KEGG) data were used for enrichment analysis of the screened differential proteins. Results: After mass spectrometry and database searching, 66 differentially expressed protein spots were identified between the UA group and the control group. Most differentially expressed proteins functioned in cytoskeleton formation, energy metabolism, or signal transduction. They were mainly involved in 50 biological processes, including peroxisome proliferator-activated receptor (PPAR) signalling and fatty acid metabolism. PPARγ and PLIN1 were subject to Western blotting analysis detection; results were consistent with the Label-Free result. Conclusion: Based on an analysis of the biological information, these proteins may be associated with the incidence and progression of the femoral neck bone tissues of hyperuricaemia patients.

Background: In type 2 diabetic mouse liver, hyperglycemia, and insulin modify gene expression. Curcumin is a powerful antioxidant and antidiabetic agent that regulates the gene expression of different signaling pathways through various transcription factors. Therefore, we hypothesized that curcumin modifies the protein expression profile in the liver of diabetic db/db mice. Objective: To determine the effects of curcumin on the liver protein profile of diabetic db/db mice. Methods: db/db and Wild Type (WT) male mice were allocated in four groups, and they were fed for eight weeks. Three WT and three diabetic db/db mice received a Standard Diet (SD; WT and db/db groups, respectively); three WT and three diabetic db/db mice received a SD supplemented with 0.75% (w/w) curcumin (WT+C and db/db+C groups, respectively). Liver proteins were separated by 2D electrophoresis. Differential protein expression analysis was performed on Image- Master 2D Platinum software, and selected proteins were identified by MALDI-TOF-MS and subjected to enrichment analysis using STRING and DAVID databases. Results: Thirty-six proteins with differential expression due to diabetic background and curcumin treatment were found; these proteins participate in the metabolism of amino acids, carbohydrates, and lipids. Interestingly, the altered expression of seven proteins was prevented in the liver of the diabetic mice that received curcumin. Conclusion: Among all differentially expressed proteins, curcumin reverted the altered expression of seven proteins. Thus, although it was observed that curcumin did not affect the biochemical parameters, it does modify the expression of some liver proteins in diabetic mice.

Background: : The global outbreak of the 2019 novel Coronavirus disease (COVID-19) caused by infection with the Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2), which appeared in China at the end of 2019, signifies a major public health issue at the current time. Objective: The objective of the present study is to characterize the physicochemical properties of the SARS-CoV-2 proteins at a residues level, and to generate a “bioinformatics fingerprint” in the form of a “PIM profile” created for each sequence utilizing the Polarity Index Method (PIM), suitable for the identification of these proteins. Methods: Two different bioinformatics approaches were used to analyze sequence characteristics of these proteins at the residues level, an in-house bioinformatics system PIM, and a set of the commonly used algorithms for the prediction of protein intrinsic disorder predisposition, such as PONDR VLXT, PONDR VL3, PONDR VSL2, PONDR FIT, IUPred_short and IUPred_long. The PIM profile was generated for four SARS-CoV-2 structural proteins and compared with the corresponding profiles of the SARS-CoV-2 non-structural proteins, SARS-CoV-2 putative proteins, SARS-- CoV proteins, MERS-CoV proteins, sets of bacterial, fungal, and viral proteins, cell-penetrating peptides, and a set of intrinsically disordered proteins. We also searched for the UniProt proteins with PIM profiles similar to those of SARS-CoV-2 structural, non-structural, and putative proteins. Results: We show that SARS-CoV-2 structural, non-structural, and putative proteins are characterized by a unique PIM profile. A total of 1736 proteins were identified from the 562,253 “reviewed” proteins from the UniProt database, whose PIM profile was similar to that of the SARS-CoV-2 structural, non-structural, and putative proteins. Conclusion: The PIM profile represents an important characteristic that might be useful for the identification of proteins similar to SARS-CoV-2 proteins.

Background: In the vast variety of viruses known, there is a particular interest in those transmitted to humans and whose ability to disseminate represents a significant public health issue. Objective: The present study’s objective is to bioinformatically characterize the proteins of the two main divisions of viruses, RNA-viruses and DNA-viruses. Methods: In this work, a set of in-house computational programs was used to calculate the polarity/ charge profiles and intrinsic disorder predisposition profiles of the proteins of several groups of viruses representing both types extracted from the UniProt database. The efficiency of these computational programs was statistically verified. Results: It was found that the polarity/charge profile of the proteins is, in most cases, an efficient discriminant that allows the re-creation of the taxonomy known for both viral groups. Additionally, the entire set of “reviewed” proteins in the UniProt database was analyzed to find proteins with polarity/ charge profiles similar to those obtained for each viral group. This search revealed a substantial number of proteins with such polarity-charge profiles. Conclusion: Polarity/charge profile represents a physicochemical metric, which is easy to calculate, and which can be used to effectively identify viral groups from their protein sequences.

Background: The association of a drug with its target protein correlates to its medicinal activity and the microenvironment plays a key role in this association. The key challenge is to identify mutations which unlikely to respond to designed drugs. Objective: Hsp70 is an anti-apoptotic factor and tumor cells overexpress Hsp70 to survive against anti-cancer agents. The impact of pathogenic mutations on Hsp70 is unknown. Elucidation of these alterations is essential to understand the molecular switch mechanism. Thus, critical spots on Hsp70 Nucleotide Binding Domain (NBD) are important since mutation-driven sensitivity may be useful in designing innovative inhibitors. Methods: ATP, AMP-PNP (non-hydrolyzable analog of ATP) along with commercially available compounds VER-155008 (ATP analog and competitive inhibitor) and MKT-077 (allosteric inhibitor of ADP bound form) were docked to Hsp70 NBD structure in silico to identify critical amino acids of inhibition mechanism. Site-directed mutagenesis of the determined critical residues along with ATP hydrolysis and luciferase refolding was performed. Wild-type and mutant Hsp70s were compared to determine the effect on protein functions in the presence or absence of inhibitors. Results: This study identified three mutants that have a loss of function for Hsp70, which may alter the drug inhibition activity as oncogenic cells have multiple mutations. Conclusion: Two commercial inhibitors employed here that mimic ATP and ADP states, respectively, are not affected by these mutational perturbations and displayed effective interference for Hsp70 functions. Designing inhibitors by considering these critical residues may improve drug design and increase drug efficiency.

Introduction: Iris taochia is an endemic plant in Turkey. Iris species has many biological effects such as antibacterial, antiinflammatory, antioxidant and anticancer properties. Apoptosis is a programmed cell death and this mechanism regulates the death of cancer cells. Purpose: The aim of our work is to investigate how the Iris taochia extracts affect the apoptotic activity in the MCF7 cells. Methods: Cytotoxic dose and cell viability is determined by the MTT assay. Bad, Bax, Bcl-2, Bcl- W, Bid, Bim, Caspase 3, Caspase 8, CD40, CD40L, cIAP-2, CytoC, DR6, Fas, FasL, HSP27, HSP60, HSP70, HTRA, IGF-I, IGF-II, IGFBP-1, IGFBP-2, IGFBP-3, IGFBP-4, IGFBP-5, IGFBP-6, IGF-1sR, Livin, p21, p27, p53, SMAC, Survivin, sTNF-R1, sTNF-R2, TNF-α, TNF-β, TRAILR-1, TRAILR-2, TRAILR-3, TRAILR-4 and XIAP proteins were measured by the membrane array kit. Results: Iris taochia extracts exhibited significant cytotoxic effects on MCF7 cells and IC50 values ranging from 1.56 to 100 μg/mL. Our results indicate that MeOH extract of Iris taochia in MCF7 cells may be a regulator of cell death proteins, cell cycle and growth factors. DCM and EtOH extracts of Iris taochia have a limited effect on MCF7 cells, especially, HSPs, which play a significant role in chemoresistance, downregulating DCM and EtOH extracts of Iris taochia, whereas ligands and receptors of extrinsic apoptotic pathway are upregulated by these extracts. Conclusion: This is the first study to investigate the cytotoxic and apoptotic effect of Iris taochia extracts on MCF7 cells. Results also showed that Iris taochia reduced cell viability and induced apoptotic pathways as a potential regulator of cancer cell death.

Background: Oxidative damage contributes to the pathophysiology of schizophrenia (SZ). Redox imbalance may lead to increased lipid peroxidation, which produces toxic aldehydes like 4-hydroxynonenal (4-HNE) ultimately leading to oxidative stress. Conversely, implications of oxidative stress point towards an alteration in HNE-protein adducts and activities of enzymatic and antioxidant systems in schizophrenia. Objectives: The present study focuses on the identification of HNE-protein adducts and their related molecular consequences in schizophrenia pathology due to oxidative stress, particularly lipid peroxidation. Materials and Methods: Oxyblotting was performed on seven autopsied brain samples each from the cortex and hippocampus region of schizophrenia patients and their respective normal healthy controls. Additionally, Thiobarbituric Acid Substances (TBARS), reduced Glutathione (GSH) levels and catalase (CAT) activities associated with oxidative stress, were also estimated. Results: Obtained results indicate substantially higher levels of oxidative stress in schizophrenia patients than healthy control group represented by elevated expression of HNE-protein adducts. Interestingly, the hippocampus region of schizophrenia brain shows increased HNE protein adducts compared to cortex. An increase in catalase activity (4.8876±1.7123) whereas a decrease in antioxidant GSH levels (0.213±0.015μmol/ml) have been observed in SZ brain. Elevated TBARS levels (0.3801±0.0532ug/ml) were obtained in brain regions of SZ patients compared with their controls that reflect increased lipid peroxidation (LPO). Conclusion: We propose the role of HNE modified proteins possibly associated with the pathology of schizophrenia. Our data revealed increased lipid peroxidation as a consequence of increased TBARS production. Furthermore, altered cellular antioxidants pathways related to GSH and CAT also highlight the involvement of oxidative stress in schizophrenia pathology.

Background and Objective: Staphylococcus aureus (S. aureus) is a gram-positive bacterium and one of the major nosocomial pathogens. It has the ability to acquire resistance against almost all available classes of antibiotics; Methicillin-Resistant S. aureus (MRSA) is a well-known antibiotic-resistant pathogen. S. aureus is a globally distributed pathogen that needs in-depth epidemiological and genomic level investigation for proper treatment and prevention. Methods: To explore the genomic epidemiology of S. aureus, in-silico Multi Locus Sequence Typing (MLST) was carried out for 355 complete genomes. Diversity within the species was investigated through pan-genome analysis and a subtractive genomic approach was employed for the identification of the core immunogenic targets. Results: Epidemiological study identified 62 different sequence types (STs) of S. aureus distributed worldwide, in which ST-8, ST-5, ST-398, ST-239, and ST-30 were the most dominant STs comprising more than 50% of the isolates. The pan-genome of S. aureus is still open with 7,199 genes and there is a major contribution (80%) of MRSA strains in the S. aureus species pangenome. The core genome (2,025 genes) of S. aureus is almost stable (comprising 72% of S. aureus genome size), while accessory and unique genes (28% of S. aureus genome size) are gradually increasing. Screening of 2,025 core genes identified putative vaccine candidates. The best scoring and dominant B-cell and T-cell epitopes were predicted out of the selected potential vaccine candidate proteins with the help of a multi-step screening procedure. Conclusion: We believe that the current study will provide insight into the genetic epidemiology and diversity of S. aureus, and the predicted epitopes against the pathogen can be tested further for their immunological responses within the host and may provide both humoral and cellular immunity against the disease.