Variations In Protein Structure Research Articles

Novel insights into fouling mechanisms in anaerobic acidification membrane bioreactor: The leading role of pH value

Anaerobic acidification membrane bioreactors (AAMBRs) have recently emerged as effective systems for recovering volatile fatty acids (VFAs) from municipal wastewater. However, membrane fouling remains a critical challenge, with its underlying mechanisms not yet fully understood. Given its importance in inhibiting methanogenic bacteria and stabilizing VFAs production, this study explores the direct and indirect effects of pH on membrane fouling mechanisms in AAMBRs. The results demonstrated that compared to pH 7, the fouling potential of anaerobic sludge was significantly higher at pH levels of 5 and 10 owing to a decrease of sludge floc size and an increase of soluble microbial products (SMP) especially proteins. Analytical techniques such as confocal laser scanning microscopy (CLSM), fourier transform infrared spectroscopy (FTIR), and fluorescence excitation-emission matrix (F-EEM) analyses revealed a substantial increase in organic foulants particularly proteins on the membrane surface at pH levels of 5 and 10. Circular dichroism (CD) spectroscopy further indicated a significant change in protein secondary structure, specifically an increase in α-helix content under extreme pH conditions. Furthermore, the extended Derjaguin-Landau-Verwey-Overbeek (XDLVO) theory indicated that elevated protein concentrations enhance the adhesion of sludge and SMP to membrane surfaces. Insights from the Flory-Huggins theory demonstrated that variations in protein structure significantly contribute to increased filtration resistance within the gel layer. Both experimental data and theoretical models underscore the dual role of pH in optimizing VFAs production and managing membrane fouling, offering insights for enhancing AAMBR performance and mitigating fouling issues.

Investigating the interaction mechanisms between arachin and resveratrol: Utilizing multi-spectroscopy and computational chemistry

Arachin (ARA) and resveratrol (RES) are the primary protein and bioactive compound in peanuts and their processed products. However, the mechanism of interaction between these two substances remained unclear. To investigate protein structural changes, conformational variations, and molecular mechanisms in the interaction between them, multispectral analysis and computational chemistry methods were employed. Experimental results confirmed that RES quenched ARA's intrinsic fluorescence through static quenching, indicating their interaction. Thermodynamic analysis revealed the interaction between them was endothermic, spontaneous, and primarily hydrophobic. Molecular dynamics (MD) simulations highlighted strong affinity between RES and ARA, with key amino acids (His425, Val426, Phe405, and Phe464) facilitating their interaction. RES binding increased stability without significant protein conformational changes. The independent gradient model based on Hirshfeld partition (IGMH) validated their interaction, emphasizing van der Waals (VDW) interactions and hydrogen bonds (H-bonds) as crucial for stable binding. This research lays a theoretical foundation for potential applications of ARA-RES complex products in the food industry.

Prediction of the antigenic regions in eight RhD variants identified by computational biology.

Changes in RHD generate variations in protein structure that lead to antigenic variants. The classical model divides them into quantitative (weak and Del) and qualitative (partial D). There are two types of protein antigens: linear and conformational. Computational biology analyses the theoretical assembly of tertiary protein structures and allows us to identify the 'topological' differences between isoforms. Our aim was to determine the theoretical antigenic differences between weak RhD variants compared with normal RhD based on structural analysis using bioinformatic techniques. We analysed the variations in secondary structures and hydrophobicity of RHD*01, RHD*01W.1, W2, W3, RHD*09.03.01, RHD*09.04, RHD*11, RHD*15 and RHD*21. We then modelled the tertiary structure and calculated their probable antigenic regions, intra-protein interactions, displacement and membrane width and compared them with Rhce. The 10 proteins are similar in their secondary structure and hydrophobicity, with the main differences observed in the exofacial coils. We identified six potential antigenic regions: one that is unique to RhD (R3), one that is common to all D (R6), three that are highly variable among RhD isoforms (R1, R2 and R4), one that they share with Rhce (R5) and two that are unique to Rhce (Ra and Rbc). The alloimmunization capacity of these subjects could be explained by the variability of the antigen pattern, which is not necessarily recognized or recognized with lower intensity by the commercially available antibodies, and not because they have a lower protein concentration in the membrane.

Cyp51A mutations, protein modeling, and efflux pump gene expression reveals multifactorial complexity towards understanding Aspergillus section Nigri azole resistance mechanism

Black Aspergillus species are the most common etiological agents of otomycosis, and pulmonary aspergillosis. However, limited data is available on their antifungal susceptibility profiles and associated resistance mechanisms. Here, we determined the azole susceptibility profiles of black Aspergillus species isolated from the Indian environment and explored the potential resistance mechanisms through cyp51A gene sequencing, protein homology modeling, and expression analysis of selected genes cyp51A, cyp51B, mdr1, and mfs based on their role in imparting resistance against antifungal drugs. In this study, we have isolated a total of 161 black aspergilli isolates from 174 agricultural soil samples. Isolates had variable resistance towards medical azoles; approximately 11.80%, 3.10%, and 1.24% of isolates were resistant to itraconazole (ITC), posaconazole (POS), and voriconazole (VRC), respectively. Further, cyp51A sequence analysis showed that non-synonymous mutations were present in 20 azole-resistant Aspergillus section Nigri and 10 susceptible isolates. However, Cyp51A homology modeling indicated insignificant protein structural variations because of these mutations. Most of the isolates showed the overexpression of mdr1, and mfs genes. Hence, the study concluded that azole-resistance in section Nigri cannot be attributed exclusively to the cyp51A gene mutation or its overexpression. However, overexpression of mdr1 and mfs genes may have a potential role in drug resistance.

Red edge excitation shift spectroscopy is highly sensitive to tryptophan composition.

Red edge excitation shift (REES) spectroscopy relies on the unique emission profiles of fluorophore-solvent interactions to profile protein molecular dynamics. Recently, we reported the use of REES to compare the stability of 32 polymorphic IgG antibodies natively containing tryptophan reporter fluorophores. Here, we expand on this work to investigate the sensitivity of REES to variations in tryptophan content using a subset of IgG3 antibodies containing arginine to tryptophan polymorphisms. Structural analysis revealed that the additional tryptophan residues were situated in highly solvated environments. Subsequently, REES showed clear differences in fluorescence emission profiles when compared with the unmutated variants, thereby limiting direct comparison of their structural dynamics. These findings highlight the exquisite sensitivity of REES to minor variations in protein structure and tryptophan composition.

Molecular Evolution and Protein Structure Variation of Dkk Family.

Dkks have inhibitory effects on the Wnt signaling pathway, which is involved in the development of skin and its appendages and the regulation of hair growth. The nucleotide sequences were compared and analyzed to further investigate the relationship between the structure and function of the Dkk gene family and vertebrate epidermal hair. The analysis of the molecular evolution of the Dkk family revealed that the evolution rate of the genes changed significantly after speciation, with the Aves and Reptilia branches showing accelerated evolution. Additionally, positive selection was observed at specific sites. The tertiary structure of the protein was also predicted. The analysis of the functional divergence of the Dkk family revealed that the functional divergence coefficient of each gene was greater than 0, with most of the functional divergence sites were located in the Cys-2 domain and a few in the Cys-1 domain. This suggests that the amino acid and functional divergence sites may play a role in regulating the binding of the Dkk family to LRP5/6, and thus affect the inhibition of Wnt signaling, leading to different functions of Dkk1, Dkk2, and Dkk4 in the development of skin hair follicles. In addition, the Dkk families of Aves and Reptilia may have undergone adaptive evolution and functional divergence.

Identification of novel SNPs in Pun1 locus for pungency in Capsicum species.

Capsaicin and its analogues known as capsaicinoids are the principal sources of pungency in Capsicum spp. In this study, characterization of North-West Himalayan chilli germplasm and commercial landraces of different Indian states known for different pungency-color combinations was done based on capsaicin concentration. Moreover, molecular variation in pungency among high, medium and mild/not pungent Capsicum spp., especially those adapted to North-West Himalayas were elucidated. Forty-nine genotypes of chilli comprising breeding lines of Kashmiri origin, commercial landraces of Southern Indian origin and one of the world's hottest chilli Bhut Jolokia from Nagaland state of India were used as an experimental material. Wide variation in capsaicin content was observed among the genotypes, wherein, Bhut Jolokia (Capsicum chinense) expressed the highest capsaicin content (10,500.75µg/g). Further, molecular analysis of PunI gene was done for discovering SNPs responsible for variations in pungency. In the non-pungent Nishat-1 (Capsicum annuum var. grossum), the 650bp DNA fragment was not amplified due to 2.5kb deletion spanning the putative promoter and first exon of AT3. The amplified DNA product for high and medium pungent was sequencing. Sequence alignment among revealed SNPs which were further observed responsible for variations in amino acid sequence and protein structure. The observed variation in protein structure might be responsible for high capsaicin production in one genotype as compared to the other and hence the protein conformation determines its interaction with the substrate.

Progress and challenges in production of recombinant Newcastle disease virus hemagglutinin-neuraminidase subunit vaccine

Recombinant Newcastle disease (ND) subunit vaccines are safe, efficacious and compatible with DIVA strategies. Hence, are promising alternatives to conventional live and inactivated vaccines for ND control and prevention. While production of hemagglutinin-neuraminidase (HN)-based ND subunit vaccines have been extensively evaluated at lab-scale, significant barriers impeding successful translation from high-cost, small bench scale production to low-cost mass production remain. This review details current state and challenges in efficacious HN subunit design, advantages and disadvantages of available production routes using different expression hosts together with gaps and solutions regarding quantitative analytical methods used for formulation in dose-efficiency studies. Critical factors influencing HN subunit vaccine immunogenicity such as protein structure, glycosylation and amino acid variations are discussed. Construct design and available expression hosts are then analyzed from viewpoint of yield and quality measured in terms of efficacy. The current deficiency in available quantitative methods for effective dose-efficacy studies is given emphasis, underscoring the importance of robust analytical methods for determination of optimal vaccine dose. The review outlines current state and way forward for the development of HN ND subunit vaccines and other recombinant subunit vaccines for veterinary applications.

Natural variations of chlorophyll fluorescence and ion transporter genes influenced the differential response of japonica rice germplasm with different salt tolerances.

Soil salinity seriously restricts rice growth, development, and production globally. Chlorophyll fluorescence and ion content reflect the level of injury and resistance of rice under salt stress. To understand the differences in the response mechanisms of japonica rice with varying degrees of salt tolerance, we analyzed the chlorophyll fluorescence characteristics and ion homeostasis of 12 japonica rice germplasm accessions by comprehensive evaluation of phenotype, haplotype, and expression of salt tolerance-related genes. The results revealed that salt-sensitive accessions were rapidly affected by the damage due to salinity. Salt tolerance score (STS) and relative chlorophyll relative content (RSPAD) were extremely significantly reduced (p<0.01), and chlorophyll fluorescence and ion homeostasis were influenced by various degrees under salt stress. The STS, RSPAD, and five chlorophyll fluorescence parameters of salt-tolerant accessions (STA) were significantly higher than that of salt-sensitive accessions (SSA). Principal component analysis (PCA) with 13 indices suggested three principal components (PCs), with a cumulative contribution rate of 90.254%, which were used to screen Huangluo (typical salt-tolerant germplasm) and Shanfuliya (typical salt-sensitive germplasm) based on the comprehensive evaluation D-value (DCI ). The expression characteristics of chlorophyll fluorescence genes (OsABCI7 and OsHCF222) and ion transporter protein genes (OsHKT1;5, OsHKT2;1, OsHAK21, OsAKT2, OsNHX1, and OsSOS1) were analyzed. The expressions of these genes were higher in Huangluo than in Shanfuliya under salt stress. Haplotype analysis revealed four key variations associated with salt tolerance, including an SNP (+1605 bp) within OsABCI7 exon, an SSR (-1231 bp) within OsHAK21 promoter, an indel site at OsNHX1 promoter (-822 bp), and an SNP (-1866 bp) within OsAKT2 promoter. Variation in OsABCI7 protein structure and differential expression of these three ion-transporter genes may contribute to the differential response of japonica rice to salt stress.

Evolution of tetraspanin antigens in the zoonotic Asian blood fluke Schistosoma japonicum

BackgroundDespite successful control efforts in China over the past 60 years, zoonotic schistosomiasis caused by Schistosoma japonicum remains a threat with transmission ongoing and the risk of localised resurgences prompting calls for a novel integrated control strategy, with an anti-schistosome vaccine as a core element. Anti-schistosome vaccine development and immunisation attempts in non-human mammalian host species, intended to interrupt transmission, and utilising various antigen targets, have yielded mixed success, with some studies highlighting variation in schistosome antigen coding genes (ACGs) as possible confounders of vaccine efficacy. Thus, robust selection of target ACGs, including assessment of their genetic diversity and antigenic variability, is paramount. Tetraspanins (TSPs), a family of tegument-surface antigens in schistosomes, interact directly with the host’s immune system and are promising vaccine candidates. Here, for the first time to our knowledge, diversity in S. japonicum TSPs (SjTSPs) and the impact of diversifying selection and sequence variation on immunogenicity in these protiens were evaluated.MethodsSjTSP sequences, representing parasite populations from seven provinces across China, were gathered by baiting published short-read NGS data and were analysed using in silico methods to measure sequence variation and selection pressures and predict the impact of selection on variation in antigen protein structure, function and antigenic propensity.ResultsHere, 27 SjTSPs were identified across three subfamilies, highlighting the diversity of TSPs in S. japonicum. Considerable variation was demonstrated for several SjTSPs between geographical regions/provinces, revealing that episodic, diversifying positive selection pressures promote amino acid variation/variability in the large extracellular loop (LEL) domain of certain SjTSPs. Accumulating polymorphisms in the LEL domain of SjTSP-2, -8 and -23 led to altered structural, functional and antibody binding characteristics, which are predicted to impact antibody recognition and possibly blunt the host’s ability to respond to infection. Such changes, therefore, appear to represent a mechanism utilised by S. japonicum to evade the host’s immune system.ConclusionWhilst the genetic and antigenic geographic variability observed amongst certain SjTSPs could present challenges to vaccine development, here we demonstrate conservation amongst SjTSP-1, -13 and -14, revealing their likely improved utility as efficacious vaccine candidates. Importantly, our data highlight that robust evaluation of vaccine target variability in natural parasite populations should be a prerequisite for anti-schistosome vaccine development.Graphical abstract

Genome wide association analysis for grain micronutrients and anti-nutritional traits in mungbean [Vigna radiata (L.) R. Wilczek] using SNP markers.

Mungbean is an important food grain legume for human nutrition and nutritional food due to its nutrient-dense seed, liked palatability, and high digestibility. However, anti-nutritional factors pose a significant risk to improving nutritional quality for bio-fortification. In the present study, genetic architecture of grain micronutrients (grain iron and zinc concentration) and anti-nutritional factors (grain phytic acid and tannin content) in association mapping panel of 145 diverse mungbean were evaluated. Based on all four parameters genotypes PUSA 1333 and IPM 02-19 were observed as desired genotypes as they had high grain iron and zinc concentration but low grain phytic acid and tannin content. The next generation sequencing (NGS)-based genotyping by sequencing (GBS) identified 14,447 genome-wide SNPs in a diverse selected panel of 127 mungbean genotypes. Population admixture analysis revealed the presence of four different ancestries among the genotypes and LD decay of ∼57.6 kb kb physical distance was noted in mungbean chromosomes. Association mapping analysis revealed that a total of 20 significant SNPs were shared by both GLM and Blink models associated with grain micronutrient and anti-nutritional factor traits, with Blink model identifying 35 putative SNPs. Further, this study identified the 185 putative candidate genes. Including potential candidate genes Vradi07g30190, Vradi01g09630, and Vradi09g05450 were found to be associated with grain iron concentration, Vradi10g04830 with grain zinc concentration, Vradi08g09870 and Vradi01g11110 with grain phytic acid content and Vradi04g11580 and Vradi06g15090 with grain tannin content. Moreover, two genes Vradi07g15310 and Vradi09g05480 showed significant variation in protein structure between native and mutated versions. The identified SNPs and candidate genes are potential powerful tools to provide the essential information for genetic studies and marker-assisted breeding program for nutritional improvement in mungbean.

Characterizing protein conformational ensembles and dynamics with 2D IR spectroscopy.

Advances in protein two-dimensional infrared spectroscopy and computational spectroscopy of amide I vibrations have provided a way to characterize the conformational variation and disorder in protein structure. These techniques use isotope-edited amide I spectra in combination with spectra computed from Markov state models built from molecular dynamics simulations to infer variation in local configuration and to refine ensembles for peptides and proteins. This talk will describe these tools drawing on examples from studies of insulin monomer and dimer structure and the dynamic interactions in the monomer-dimer equilibrium.

Genetic and functional odorant receptor variation in the Homo lineage

Humans, Neanderthals, and Denisovans independently adapted to a wide range of geographic environments and their associated food odors. Using ancient DNA sequences, we explored the invitro function of thirty odorant receptor genes in the genus Homo. Our extinct relatives had highly conserved olfactory receptor sequence, but humans did not. Variations in odorant receptor protein sequence and structure may have produced variation in odor detection and perception. Variants led to minimal changes in specificity but had more influence on functional sensitivity. The few Neanderthal variants disturbed function, whereas Denisovan variants increased sensitivity to sweet and sulfur odors. Geographic adaptations may have produced greater functional variation in our lineage, increasing our olfactory repertoire and expanding our adaptive capacity. Our survey of olfactory genes and odorant receptors suggests that our genus has a shared repertoire with possible local ecological adaptations.

Mechanism of the reduced allergenicity of shrimp (Macrobrachium nipponense) by combined thermal/pressure processing: Insight into variations in protein structure, gastrointestinal digestion and immunodominant linear epitopes

This study aimed to explore the underlying mechanism about combined thermal/pressure processing on the allergenicity of shrimp (Macrobrachium nipponense). We analysed sensitizing and eliciting capacities, structural changes, gastrointestinal digestion, and mapped linear epitopes. Mice treated with steamed + reverse-pressure sterilized shrimp exhibited lower specific IgE and IgG1 concentrations, degranulation, vascular permeability, and allergic symptoms than those fed with raw shrimp or steamed shrimp (p < 0.05). Reduced allergenicity of shrimp using combined thermal/pressure processing was not only associated with protein unfolding and exposure of hydrophobic residues, but also related to disruption of immunodominant linear epitopes (Glu177-Ser188 in tropomyosin, Gln361-Ser366 in β-actin) due to changes in gastrointestinal digestion behavior. Moreover, heat/digested stable epitopes of arginine kinase were located inside its 3D structure, preventing binding with IgE and maintaining hypoallergenicity following combined processing. Thus, steaming and reverse-pressure sterilization might be an efficient low-allergenic food processing method for Macrobrachium nipponense.

In vitro inhibition of glucose gastro-intestinal enzymes and antioxidant activity of hydrolyzed collagen peptides from different species.

The growing value of industrial collagen by-products has given rise to interest in extracting them from different species of animals. Intrinsic protein structure variation of collagen sources and its hydrolysis can bring about different bioactivities. This study aimed to characterize and evaluate the differences in vitro biological potential of commercial bovine (BH), fish (FH), and porcine hydrolysates (PH) regarding their antioxidant and hypoglycemic activities. All samples showed percentages above 90% of protein content, with high levels of amino acids (glycine, proline, and hydroxyproline), responsible for the specific structure of collagen. The BH sample showed a higher degree of hydrolysis (DH) (8.7%) and a higher percentage of smaller than 2kDa peptides (74.1%). All collagens analyzed in vitro showed inhibition of pancreatic enzymes (α-amylase and α-glucosidase), with the potential to prevent diabetes mellitus. The PH sample showed higher antioxidant activities measured by ORAC (67.08 ± 4.23 μmol Trolox Eq./g) and ABTS radical scavenging (65.69 ± 3.53 μmol Trolox Eq./g) methods. For the first time, DNA protection was analyzed to hydrolyzed collagen peptides, and the FH sample showed a protective antioxidant action to supercoiled DNA both in the presence (39.51%) and in the absence (96.36%) of AAPH (reagent 2,2'-azobis(2-amidinopropane)). The results confirmed that the source of native collagen reflects on the bioactivity of hydrolyzed collagen peptides, probably due to its amino acid composition. PRACTICAL APPLICATIONS: Our data provide new application for collagen hydrolysates with hypoglycemiant and antioxidant activity. These data open discussion for future studies on the additional benefits arising from collagen peptide consumption for the prevention of aging complications or hyperglycemic conditions as observed in chronic diseases such as diabetes mellitus type II (DM 2). The confirmation of these results can open new market areas for the use of collagen with pharmacological applications or to produce new supplements. Furthermore, provides a solution for waste collagen from meat industries and adds value to the product.

Amino acid transporter (AAT) gene family in Tartary buckwheat (Fagopyrum tataricum L. Gaertn.): Characterization, expression analysis and functional prediction

Tartary buckwheat (Fagopyrum tataricum L. Gaertn., TB) is an ancient minor crop and an important food source for humans to supplement nutrients such as flavonoids and essential amino acids. Amino acid transporters (AATs) play critical roles in plant growth and development through the transport of amino acids. In this study, 104 AATs were identified in TB genome and divided into 11 subfamilies by phylogenetic relationships. Tandem and segmental duplications promoted the expansion of FtAAT gene family, and the variations of gene sequence, protein structure and expression pattern were the main reasons for the functional differentiation of FtAATs. Based on RNA-seq and qRT-PCR, the expression patterns of FtAATs in different tissues and under different abiotic stresses were analyzed, and several candidate FtAATs that might affect grain development and response to abiotic stresses were identified, such as FtAAP12 and FtCAT7. Finally, combined with the previous studies, the expression patterns and phylogenetic relationships of AATs in multiple species, the functions of multiple high-confidence FtAAT genes were predicted, and the schematic diagram of FtAATs in TB was initially drawn. Overall, this work provided a framework for further functional analysis of FtAAT genes and important clues for the improvement of TB quality and stress resistance.

PREDICTED PROTEIN STRUCTURE VARIATIONS INDICATE THE CLINICAL PRESENTATION OF CYP4V2-RELATED BIETTI CRYSTALLINE DYSTROPHY.

To investigate the relationship between different CYP4V2 disease-causing variants and disease severity in Bietti crystalline dystrophy (BCD). Twenty-one subjects from 19 unrelated families with a clinical diagnosis of BCD were enrolled. A novel severity prediction score for BCD based on the predicted molecular impact of CYP4V2 variants was applied for grouping and subsequent analyses. The more severe variants led to less CYP4V2 protein function preservation and a higher severity prediction score. All subjects harbored two alleles of CYP4V2 disease-causing variants, of which c.802-8_810del17insGC was the most prevalent (14/21, 66.67%) and c.1507G>C was novel. According to the severity score, the subjects were categorized into severe, moderate, and mild groups with different preservation of central vision (mean logMAR visual acuity 0.95 ± 0.82, 0.89 ± 1.22, and 0.56 ± 0.64, respectively). The patients with a lower severity score had slower disease progression. This is the first cohort study of BCD in Taiwan, and we established a novel BCD severity index based on the molecular impact of different CYP4V2 variants. More severe impairment of CYP4V2 protein led to a more severe disease course with earlier progression. Our results could be helpful in identifying a therapeutic window for patients with BCD.

A Glance into MTHFR Deficiency at a Molecular Level.

MTHFR deficiency still deserves an investigation to associate the phenotype to protein structure variations. To this aim, considering the MTHFR wild type protein structure, with a catalytic and a regulatory domain and taking advantage of state-of-the-art computational tools, we explore the properties of 72 missense variations known to be disease associated. By computing the thermodynamic ΔΔG change according to a consensus method that we recently introduced, we find that 61% of the disease-related variations destabilize the protein, are present both in the catalytic and regulatory domain and correspond to known biochemical deficiencies. The propensity of solvent accessible residues to be involved in protein-protein interaction sites indicates that most of the interacting residues are located in the regulatory domain, and that only three of them, located at the interface of the functional protein homodimer, are both disease-related and destabilizing. Finally, we compute the protein architecture with Hidden Markov Models, one from Pfam for the catalytic domain and the second computed in house for the regulatory domain. We show that patterns of disease-associated, physicochemical variation types, both in the catalytic and regulatory domains, are unique for the MTHFR deficiency when mapped into the protein architecture.

RETRACTED ARTICLE: Deterministic functions for measuring human protein structural variations with merit based ensemble learning scheme for native classification

RETRACTED ARTICLE: Deterministic functions for measuring human protein structural variations with merit based ensemble learning scheme for native classification

In silico mechanistic model of microtubule assembly inhibition by selective chromone derivatives

Microtubules and actin filaments are cytoskeletal protein assemblies that are engaged in motility, mitosis, and regulate cell signalling with an essential role in the control of protein synthesis and accordingly, cell growth. Inhibition of cytoskeletal organelles, in particular tubulin polymerization, is important in cancer therapy due to their antimitotic role.Considering protein structure variations, we conducted a molecular docking study followed by a series of molecular dynamics (MD) simulations at different binding sites to investigate how a known α-/β-tubulin dimerization inhibitor, Glaziovianin A, and its analogue Gatastatin target human γ-tubulin subunit. The results of our MD simulations reproduced these protein-ligand interactions, demonstrating that Glaziovianin A is selective against β-tubulin's colchicine site and Gatastatin hits γ-tubulin's GTP binding site.Further analyses shed light on the reasons for subunit-focused selectivity for these structurally similar compounds, provided mechanistic insights into some of the tubulin functions such as the conformational transitions and subsequent microtubule disruption. The information combined from energy and structural analysis of MD, with all represented validation steps, and detailed study of specific protein-protein interaction features potentially could facilitate the early identification of γ-tubulin-specific inhibitors.