Displaying publications 1 - 20 of 27 in total

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  1. Fulltext Molten globule-like partially folded state of Bacillus licheniformis α-amylase at low pH induced by 1,1,1,3,3,3-hexafluoroisopropanol
    Abd Halim AA, Zaroog MS, Kadir HA, Tayyab S
    ScientificWorldJournal, 2014;2014:824768.
    PMID: 24977228 DOI: 10.1155/2014/824768
    Effect of 1,1,1,3,3,3-hexafluoroisopropanol (HFIP) on acid-denatured Bacillus licheniformis α -amylase (BLA) at pH 2.0 was investigated by far-UV CD, intrinsic fluorescence, and ANS fluorescence measurements. Addition of increasing HFIP concentrations led to an increase in the mean residue ellipticity at 222 nm (MRE 222 nm) up to 1.5 M HFIP concentration beyond which it sloped off. A small increase in the intrinsic fluorescence and a marked increase in the ANS fluorescence were also observed up to 0.4 M HFIP concentration, both of which decreased thereafter. Far- and near-UV CD spectra of the HFIP-induced state observed at 0.4 M HFIP showed significant retention of the secondary structures closer to native BLA but a disordered tertiary structure. Increase in the ANS fluorescence intensity was also observed with the HFIP-induced state, suggesting exposure of the hydrophobic clusters to the solvent. Furthermore, thermal denaturation of HFIP-induced state showed a non-cooperative transition. Taken together, all these results suggested that HFIP-induced state of BLA represented a molten globule-like state at pH 2.0.
    Matched MeSH terms: Protein Folding
  2. Deciphering the flexibility and dynamics of Geobacillus zalihae strain T1 lipase at high temperatures by molecular dynamics simulation
    Abdul Rahman MB, Karjiban RA, Salleh AB, Jacobs D, Basri M, Thean Chor AL, et al.
    Protein Pept Lett, 2009;16(11):1360-70.
    PMID: 20001926
    The stability of biocatalysts is an important criterion for a sustainable industrial operation economically. T1 lipase is a thermoalkalophilic enzyme derived from Geobacillus zalihae strain T1 (T1 lipase) that was isolated from palm oil mill effluent (POME) in Malaysia. We report here the results of high temperatures molecular dynamics (MD) simulations of T1 lipase in explicit solvent. We found that the N-terminal moiety of this enzyme was accompanied by a large flexibility and dynamics during temperature-induced unfolding simulations which preceded and followed by clear structural changes in two specific regions; the small domain (consisting of helices alpha3 and alpha5, strands beta1 and beta2, and connecting loops) and the main catalytic domain or core domain (consisting of helices alpha6- alpha9 and connecting loops which located above the active site) of the enzyme. The results suggest that the small domain of model enzyme is a critical region to the thermostability of this organism.
    Matched MeSH terms: Protein Folding
  3. Molecular dynamics of thermoenzymes at high temperature and pressure: a review
    Abedi Karjiban R, Lim WZ, Basri M, Abdul Rahman MB
    Protein J, 2014 Aug;33(4):369-76.
    PMID: 24871480 DOI: 10.1007/s10930-014-9568-8
    Lipases are known for their versatility in addition to their ability to digest fat. They can be used for the formulation of detergents, as food ingredients and as biocatalysts in many industrial processes. Because conventional enzymes are frangible at high temperatures, the replacement of conventional chemical routes with biochemical processes that utilize thermostable lipases is vital in the industrial setting. Recent theoretical studies on enzymes have provided numerous fundamental insights into the structures, folding mechanisms and stabilities of these proteins. The studies corroborate the experimental results and provide additional information regarding the structures that were determined experimentally. In this paper, we review the computational studies that have described how temperature affects the structure and dynamics of thermoenzymes, including the thermoalkalophilic L1 lipase derived from Bacillus stearothermophilus. We will also discuss the potential of using pressure for the analysis of the stability of thermoenzymes because high pressure is also important for the processing and preservation of foods.
    Matched MeSH terms: Protein Folding
  4. Biophysical characterization of a recombinant lipase KV1 from Acinetobacter haemolyticus in relation to pH and temperature
    Batumalaie K, Khalili E, Mahat NA, Huyop F, Wahab RA
    Biochimie, 2018 Sep;152:198-210.
    PMID: 30036604 DOI: 10.1016/j.biochi.2018.07.011
    Spectroscopic and calorimetric methods were employed to assess the stability and the folding aspect of a novel recombinant alkaline-stable lipase KV1 from Acinetobacter haemolyticus under varying pH and temperature. Data on far ultraviolet-circular dichroism of recombinant lipase KV1 under two alkaline conditions (pH 8.0 and 12.0) at 40 °C reveal strong negative ellipticities at 208, 217, 222 nm, implying its secondary structure belonging to a α + β class with 47.3 and 39.0% ellipticity, respectively. Results demonstrate that lipase KV1 adopts its most stable conformation at pH 8.0 and 40 °C. Conversely, the protein assumes a random coil structure at pH 4.0 and 80 °C, evident from a strong negative peak at ∼ 200 nm. This blue shift suggests a general decline in enzyme activity in conjunction with the partially or fully unfolded state that invariably exposed more hydrophobic surfaces of the lipase protein. The maximum emission at ∼335 nm for pH 8.0 and 40 °C indicates the adoption of a favorable protein conformation with a high number of buried tryptophan residues, reducing solvent exposure. Appearance of an intense Amide I absorption band at pH 8.0 corroborates an intact secondary structure. A lower enthalpy value for pH 4.0 over pH 8.0 and 12.0 in the differential scanning calorimetric data corroborates the stability of the lipase at alkaline conditions, while a low Km (0.68 ± 0.03 mM) for tributyrin verifies the high affinity of lipase KV1 for the substrate. The data, herein offer useful insights into future structure-based tunable catalytic activity of lipase KV1.
    Matched MeSH terms: Protein Folding
  5. Fulltext FKBP22 from the psychrophilic bacterium Shewanella sp. SIB1 selectively binds to the reduced state of insulin to prevent its aggregation
    Budiman C, Goh CKW, Arief II, Yusuf M
    Cell Stress Chaperones, 2021 Mar;26(2):377-386.
    PMID: 33247372 DOI: 10.1007/s12192-020-01183-0
    FKBP22 of a psychrophilic bacterium, Shewanella sp. SIB1 (SIB1 FKBP22), is a member of peptidyl-prolyl cis-trans isomerase (PPIase) and consists of N- and C-domains responsible for chaperone-like and PPIase catalytic activities, respectively. The chaperone-like activity of SIB1 FKBP22 was previously evidenced by its ability to prevent dithiothreitol (DTT)-induced insulin aggregation. Nevertheless, the mechanism by which this protein inhibits the aggregation remains unclear. To address this, the binding affinity of SIB1 FKBP22 to the native or reduced states of insulin was examined using surface plasmon resonance (SPR). The native and reduced states refer to insulin in the absence or DTT presence, respectively. The SPR sensorgram showed that SIB1 FKBP22 binds specifically to the reduced state of insulin, with a KD value of 37.31 ± 3.20 μM. This binding was facilitated by the N-domain, as indicated by the comparable KD values of the N-domain and SIB1 FKBP22. Meanwhile, the reduced state of insulin was found to have no affinity towards the C-domain. The KD value of SIB1 FKBP22 was slightly decreased by NaCl but was not severely affected by FK506, a specific FKBP inhibitor. Similarly, the prevention of DTT-induced aggregation by SIB1 FKBP22 was also modulated by the N-domain and was not affected by FK506. Further, the reduced and native states of insulin had no effect on the catalytic efficiency (kcat/KM) of SIB1 FKBP22 towards a peptide substrate. Nevertheless, the reduced state of insulin slightly reduced the catalytic efficiency towards refolding RNase T1, at up to 1.5-fold lower than in the absence of insulin. These results suggested that the binding event was mainly facilitated by hydrophobic interaction and was independent from its PPIase activity. Altogether, a possible mechanism by which SIB1 FKBP22 prevents DTT-induced insulin aggregation was proposed.
    Matched MeSH terms: Protein Folding
  6. Fold prediction problem: the application of new physical and physicochemical-based features
    Dehzangi A, Phon-Amnuaisuk S
    Protein Pept Lett, 2011 Feb;18(2):174-85.
    PMID: 21054271
    One of the most important goals in bioinformatics is the ability to predict tertiary structure of a protein from its amino acid sequence. In this paper, new feature groups based on the physical and physicochemical properties of amino acids (size of the amino acids' side chains, predicted secondary structure based on normalized frequency of β-Strands, Turns, and Reverse Turns) are proposed to tackle this task. The proposed features are extracted using a modified feature extraction method adapted from Dubchak et al. To study the effectiveness of the proposed features and the modified feature extraction method, AdaBoost.M1, Multi Layer Perceptron (MLP), and Support Vector Machine (SVM) that have been commonly and successfully applied to the protein folding problem are employed. Our experimental results show that the new feature groups altogether with the modified feature extraction method are capable of enhancing the protein fold prediction accuracy better than the previous works found in the literature.
    Matched MeSH terms: Protein Folding*
  7. Does recovery in the spectral characteristics of GdnHCl-denatured Bacillus licheniformis α-amylase due to added calcium point towards protein stabilization?
    Halim AA, Feroz SR, Tayyab S
    Biosci Biotechnol Biochem, 2013;77(1):87-96.
    PMID: 23291750
    Treatment of Bacillus licheniformis α-amylase (BLA) with guanidine hydrochloride (GdnHCl) produced both denatured and aggregated forms of the enzyme as studied by circular dichroism, fluorescence, UV difference spectroscopy, size exclusion chromatography (SEC), and enzymatic activity. The presence of CaCl(2) in the incubation mixture produced significant recovery in spectral signals, being complete in presence of 10 mM CaCl(2), as well as in enzymatic activity, which is indicative of protein stabilization. However, the SEC results obtained with GdnHCl-denatured BLA both in the absence and the presence of 10 mM CaCl(2) suggested significant aggregation of the protein in the absence of CaCl(2) and disaggregation in its presence. Although partial structural stabilization with significant retention of enzymatic activity was observed in the presence of calcium, it was far from the native state, as reflected by spectral probes. Hence, spectral results as to BLA stabilization should be treated with caution in the presence of aggregation.
    Matched MeSH terms: Protein Folding
  8. Contribution of process-induced molten-globule state formation in duck liver protein to the enhanced binding ability of (E,E)-2,4-heptadienal
    Han C, Zheng Y, Wang L, Zhou C, Wang J, He J, et al.
    J Sci Food Agric, 2023 May;103(7):3334-3345.
    PMID: 36786016 DOI: 10.1002/jsfa.12499
    BACKGROUND: Extracted proteins of alternative animal origin tend to present strong off-flavor perception due to physicochemical interactions of coextracted off-flavor compounds with proteins. To investigate the relationship between absorption behaviors of volatile aromas and the processes-induced variations in protein microstructures and molecular conformations, duck liver protein isolate (DLp) was subjected to heating (65/100 °C, 15 min) and ultra-high pressure (UHP, 100-500 MPa/10 min, 28 °C) treatments to obtain differential unfolded protein states.

    RESULTS: Heat and UHP treatments induced the unfolding of DLp to varied degrees, as revealed by fluorescence spectroscopy, ultraviolet-visible absorption, circular dichroism spectra and surface hydrophobicity measurements. Two types of heating-denatured states with varied unfolding degrees were obtained, while UHP at both levels of 100/500 MPa caused partial unfolding of DLp and the presence of a molten-globule state, which significantly enhanced the binding affinity between DLp and (E,E)-2,4-heptadienal. In particular, significantly modified secondary structures of DLp were observed in heating-denatured samples. Excessive denaturing and unfolding degrees resulted in no significant changes in the absorption behavior of the volatile ligand, as characterized by observations of fluorescence quenching and analysis of headspace concentrations.

    CONCLUSION: Defining process-induced conformational transition behavior of matrix proteins could be a promising strategy to regulate food flavor attributes and, particularly, to produce DLp coextracted with limited off-flavor components by modifying their interaction during extraction processes. © 2023 Society of Chemical Industry.

    Matched MeSH terms: Protein Folding*
  9. Conformational analysis of champedak galactose-binding lectin under different urea concentrations
    Kameel NI, Wong YH, Shuib AS, Tayyab S
    Plant Physiol Biochem, 2016 Jan;98:57-63.
    PMID: 26642433 DOI: 10.1016/j.plaphy.2015.11.007
    Conformational analysis of champedak galactose-binding (CGB) lectin under different urea concentrations was studied in phosphate-buffered saline (pH 7.2) using far-ultraviolet circular dichroism (far-UV CD), tryptophan (Trp) fluorescence and ANS fluorescence. In all cases, CGB lectin displayed a two-step, three-state transition. The first transition (from the native state to the intermediate state) started at ∼2.0 M urea and ended at ∼4.5 M urea, while the second transition (from the intermediate state to the completely denatured state) was characterized by the start- and end-points at ∼5.75 M and ∼7.5 M urea, respectively, when analyzed by the emission maximum of Trp fluorescence. A marked increase in the Trp fluorescence, ANS fluorescence and -CD values at 218 nm (-CD218 nm) represented the first transition, whereas a decrease in these parameters defined the second transition. On the other hand, emission maximum of the Trp fluorescence showed a continuous increase throughout the urea concentration range. Transformation of tetramer into monomer represented the first transition, whereas the second transition reflected the unfolding of monomer. Far-UV CD, Trp fluorescence and ANS fluorescence spectra were used to characterize the native, the intermediate and the completely denatured states of CGB lectin, obtained at 0.0 M, 5.0 M and 9.0 M urea, respectively. The intermediate state was characterized by the presence of higher secondary structures, increased ANS binding as well as increased Trp fluorescence intensity. A gradual decrease in the hemagglutination activity of CGB lectin was observed with increasing urea concentrations, showing complete loss at 4.0 M urea.
    Matched MeSH terms: Protein Folding/drug effects
  10. Fulltext Intracellular Targeting Mechanisms by Antimicrobial Peptides
    Le CF, Fang CM, Sekaran SD
    Antimicrob Agents Chemother, 2017 04;61(4).
    PMID: 28167546 DOI: 10.1128/AAC.02340-16
    Antimicrobial peptides (AMPs) are expressed in various living organisms as first-line host defenses against potential harmful encounters in their surroundings. AMPs are short polycationic peptides exhibiting various antimicrobial activities. The principal antibacterial activity is attributed to the membrane-lytic mechanism which directly interferes with the integrity of the bacterial cell membrane and cell wall. In addition, a number of AMPs form a transmembrane channel in the membrane by self-aggregation or polymerization, leading to cytoplasm leakage and cell death. However, an increasing body of evidence has demonstrated that AMPs are able to exert intracellular inhibitory activities as the primary or supportive mechanisms to achieve efficient killing. In this review, we focus on the major intracellular targeting activities reported in AMPs, which include nucleic acids and protein biosynthesis and protein-folding, protease, cell division, cell wall biosynthesis, and lipopolysaccharide inhibition. These multifunctional AMPs could serve as the potential lead peptides for the future development of novel antibacterial agents with improved therapeutic profiles.
    Matched MeSH terms: Protein Folding/drug effects
  11. Expression of a codon-optimised recombinant Ara h 2.02 peanut allergen in Escherichia coli
    Lew MH, Lim RL
    Appl Microbiol Biotechnol, 2016 Jan;100(2):661-71.
    PMID: 26411458 DOI: 10.1007/s00253-015-6953-y
    Current diagnostic tools for peanut allergy using crude peanut extract showed low predictive value and reduced specificity for detection of peanut allergen-specific immunoglobulin E (IgE). The Ara h 2.02, an isoform of the major peanut allergen Ara h 2, contains three IgE epitope recognition sequence of 'DPYSPS' and may be a better reagent for component resolve diagnosis. This research aimed to generate a codon-optimised Ara h 2.02 gene for heterologous expression in Escherichia coli and allergenicity study of this recombinant protein. The codon-optimised gene was generated by PCR using overlapping primers and cloned into the pET-28a (+) expression vector. Moderate expression of a 22.5 kDa 6xhistidine-tagged recombinant Ara h 2.02 protein (6xHis-rAra h 2.02) in BL21 (DE3) host cells was observed upon induction with 1 mM isopropyl β-D-1-thiogalactopyranoside (IPTG). The insoluble recombinant protein was purified under denaturing condition using nickel-nitrilotriacetic acid (Ni-NTA) affinity chromatography and refolded by dialysis in decreasing urea concentration, amounting to a yield of 74 mg/l of expression culture. Matrix-assisted laser desorption/ionisation time-of-flight (MALDI-TOF) and immunoblot analysis confirmed the production of the recombinant 6xHis-rAra h 2.02. The refolded recombinant 6xHis-rAra h 2.02, with or without adjuvant, was able to elicit comparable level of allergen-specific IgE and IgG1 in sensitised Balb/c mice. In addition, the specific IgE antibodies raised against the recombinant protein were able to recognise the native Ara h 2 protein, demonstrating its allergenicity and potential as a reagent for diagnosis and therapeutic study.
    Matched MeSH terms: Protein Folding
  12. Fulltext IntFOLD: an integrated web resource for high performance protein structure and function prediction
    McGuffin LJ, Adiyaman R, Maghrabi AHA, Shuid AN, Brackenridge DA, Nealon JO, et al.
    Nucleic Acids Res, 2019 07 02;47(W1):W408-W413.
    PMID: 31045208 DOI: 10.1093/nar/gkz322
    The IntFOLD server provides a unified resource for the automated prediction of: protein tertiary structures with built-in estimates of model accuracy (EMA), protein structural domain boundaries, natively unstructured or disordered regions in proteins, and protein-ligand interactions. The component methods have been independently evaluated via the successive blind CASP experiments and the continual CAMEO benchmarking project. The IntFOLD server has established its ranking as one of the best performing publicly available servers, based on independent official evaluation metrics. Here, we describe significant updates to the server back end, where we have focused on performance improvements in tertiary structure predictions, in terms of global 3D model quality and accuracy self-estimates (ASE), which we achieve using our newly improved ModFOLD7_rank algorithm. We also report on various upgrades to the front end including: a streamlined submission process, enhanced visualization of models, new confidence scores for ranking, and links for accessing all annotated model data. Furthermore, we now include an option for users to submit selected models for further refinement via convenient push buttons. The IntFOLD server is freely available at: http://www.reading.ac.uk/bioinf/IntFOLD/.
    Matched MeSH terms: Protein Folding
  13. Remote protein homology detection and fold recognition using two-layer support vector machine classifiers
    Muda HM, Saad P, Othman RM
    Comput Biol Med, 2011 Aug;41(8):687-99.
    PMID: 21704312 DOI: 10.1016/j.compbiomed.2011.06.004
    Remote protein homology detection and fold recognition refer to detection of structural homology in proteins where there are small or no similarities in the sequence. To detect protein structural classes from protein primary sequence information, homology-based methods have been developed, which can be divided to three types: discriminative classifiers, generative models for protein families and pairwise sequence comparisons. Support Vector Machines (SVM) and Neural Networks (NN) are two popular discriminative methods. Recent studies have shown that SVM has fast speed during training, more accurate and efficient compared to NN. We present a comprehensive method based on two-layer classifiers. The 1st layer is used to detect up to superfamily and family in SCOP hierarchy using optimized binary SVM classification rules. It used the kernel function known as the Bio-kernel, which incorporates the biological information in the classification process. The 2nd layer uses discriminative SVM algorithm with string kernel that will detect up to protein fold level in SCOP hierarchy. The results obtained were evaluated using mean ROC and mean MRFP and the significance of the result produced with pairwise t-test was tested. Experimental results show that our approaches significantly improve the performance of remote protein homology detection and fold recognition for all three different version SCOP datasets (1.53, 1.67 and 1.73). We achieved 4.19% improvements in term of mean ROC in SCOP 1.53, 4.75% in SCOP 1.67 and 4.03% in SCOP 1.73 datasets when compared to the result produced by well-known methods. The combination of first layer and second layer of BioSVM-2L performs well in remote homology detection and fold recognition even in three different versions of datasets.
    Matched MeSH terms: Protein Folding
  14. Fulltext Paradigm Shift in Drug Re-purposing From Phenalenone to Phenaleno-Furanone to Combat Multi-Drug Resistant Salmonella enterica Serovar Typhi
    Mujawar S, Gatherer D, Lahiri C
    Front Cell Infect Microbiol, 2018;8:402.
    PMID: 30488026 DOI: 10.3389/fcimb.2018.00402
    Over recent years, typhoid fever has gained increasing attention with several cases reporting treatment failure due to multidrug resistant (MDR) strains of Salmonella enterica serovar Typhi. While new drug development strategies are being devised to combat the threat posed by these MDR pathogens, drug repurposing or repositioning has become a good alternative. The latter is considered mainly due to its capacity for saving sufficient time and effort for pre-clinical and optimization studies. Owing to the possibility of an unsuccessful repositioning, due to the mismatch in the optimization of the drug ligand for the changed biochemical properties of "old" and "new" targets, we have chosen a "targeted" approach of adopting a combined chemical moiety-based drug repurposing. Using small molecules selected from a combination of earlier approved drugs having phenalenone and furanone moieties, we have computationally delineated a step-wise approach to drug design against MDR Salmonella. We utilized our network analysis-based pre-identified, essential chaperone protein, SicA, which regulates the folding and quality of several secretory proteins including the Hsp70 chaperone, SigE. To this end, another crucial chaperone protein, Hsp70 DnaK, was also considered due to its importance for pathogen survival under the stress conditions typically encountered during antibiotic therapies. These were docked with the 19 marketed anti-typhoid drugs along with two phenalenone-furanone derivatives, 15 non-related drugs which showed 70% similarity to phenalenone and furanone derivatives and other analogous small molecules. Furthermore, molecular dynamics simulation studies were performed to check the stability of the protein-drug complexes. Our results showed the best binding interaction and stability, under the parameters of a virtual human body environment, with XR770, a phenaleno-furanone moiety based derivative. We therefore propose XR770, for repurposing for therapeutic intervention against emerging and significant drug resistance conferred by pathogenic Salmonella strains.
    Matched MeSH terms: Protein Folding/drug effects
  15. Fulltext Crystallization and preliminary X-ray analysis of recombinant Glomerella cingulata cutinase
    Nyon MP, Rice DW, Berrisford JM, Huang H, Moir AJ, Craven CJ, et al.
    Acta Crystallogr Sect F Struct Biol Cryst Commun, 2008 Jun 01;64(Pt 6):504-8.
    PMID: 18540061 DOI: 10.1107/S1744309108012086
    Cutinase catalyzes the hydrolysis of water-soluble esters and long-chain triglycerides and belongs to the family of serine hydrolases. The enzyme is thought to represent an evolutionary link between the esterase and lipase families and has potential applications in a wide range of industrial hydrolytic processes, for which an understanding of the molecular basis of its substrate specificity is critical. Glomerella cingulata cutinase has been cloned and the protein has been overexpressed in Escherichia coli, purified and subsequently crystallized in a wide range of different crystal forms in the presence and absence of inhibitors. The best crystals are those of the apo cutinase, which diffract to beyond 1.6 A resolution and belong to space group P4(1)2(1)2 or P4(3)2(1)2. Crystals of cutinase with the inhibitors PETFP or E600 belong to space groups P2(1)2(1)2(1) and P2(1), respectively, and diffract to approximately 2.5 A resolution. All of the crystals are suitable for structural studies, which are currently ongoing.
    Matched MeSH terms: Protein Folding
  16. Fulltext Characterisation of a New Fungal Immunomodulatory Protein from Tiger Milk mushroom, Lignosus rhinocerotis
    Pushparajah V, Fatima A, Chong CH, Gambule TZ, Chan CJ, Ng ST, et al.
    Sci Rep, 2016 07 27;6:30010.
    PMID: 27460640 DOI: 10.1038/srep30010
    Lignosus rhinocerotis (Tiger milk mushroom) is an important folk medicine for indigenous peoples in Southeast Asia. We previously reported its de novo assembled 34.3 Mb genome encoding a repertoire of proteins including a putative bioactive fungal immunomodulatory protein. Here we report the cDNA of this new member (FIP-Lrh) with a homology range of 54-64% to FIPs from other mushroom species, the closest is with FIP-glu (LZ-8) (64%) from Ganoderma lucidum. The FIP-Lrh of 112 amino acids (12.59 kDa) has a relatively hydrophobic N-terminal. Its predicted 3-dimensional model has identical folding patterns to FIP-fve and contains a partially conserved and more positively charged carbohydrates binding pocket. Docking predictions of FIP-Lrh on 14 glycans commonly found on cellular surfaces showed the best binding energy of -3.98 kcal/mol to N-acetylgalactosamine and N-acetylglucosamine. Overexpression of a 14.9 kDa soluble 6xHisFIP-Lrh was achieved in pET-28a(+)/BL21 and the purified recombinant protein was sequence verified by LC-MS/MS (QTOF) analysis. The ability to haemagglutinate both mouse and human blood at concentration ≥0.34 μM, further demonstrated its lectin nature. In addition, the cytotoxic effect of 6xHisFIP-Lrh on MCF-7, HeLa and A549 cancer cell lines was detected at IC50 of 0.34 μM, 0.58 μM and 0.60 μM, respectively.
    Matched MeSH terms: Protein Folding
  17. Fulltext The structure of a major surface antigen SAG19 from Eimeria tenella unifies the Eimeria SAG family
    Ramly NZ, Dix SR, Ruzheinikov SN, Sedelnikova SE, Baker PJ, Chow YP, et al.
    Commun Biol, 2021 03 19;4(1):376.
    PMID: 33742128 DOI: 10.1038/s42003-021-01904-w
    In infections by apicomplexan parasites including Plasmodium, Toxoplasma gondii, and Eimeria, host interactions are mediated by proteins including families of membrane-anchored cysteine-rich surface antigens (SAGs) and SAG-related sequences (SRS). Eimeria tenella causes caecal coccidiosis in chickens and has a SAG family with over 80 members making up 1% of the proteome. We have solved the structure of a representative E. tenella SAG, EtSAG19, revealing that, despite a low level of sequence similarity, the entire Eimeria SAG family is unified by its three-layer αβα fold which is related to that of the CAP superfamily. Furthermore, sequence comparisons show that the Eimeria SAG fold is conserved in surface antigens of the human coccidial parasite Cyclospora cayetanensis but this fold is unrelated to that of the SAGs/SRS proteins expressed in other apicomplexans including Plasmodium species and the cyst-forming coccidia Toxoplasma gondii, Neospora caninum and Besnoitia besnoiti. However, despite having very different structures, Consurf analysis showed that Eimeria SAG and Toxoplasma SRS families each exhibit marked hotspots of sequence hypervariability that map to their surfaces distal to the membrane anchor. This suggests that the primary and convergent purpose of the different structures is to provide a platform onto which sequence variability can be imposed.
    Matched MeSH terms: Protein Folding
  18. Construction of A Preliminary Three-Dimensional Structure Simian betaretrovirus Serotype-2 (SRV-2) Reverse Transcriptase Isolated from Indonesian Cynomolgus Monkey
    Saepuloh U, Iskandriati D, Pamungkas J, Solihin DD, Mariya SS, Sajuthi D
    Trop Life Sci Res, 2020 Oct;31(3):47-61.
    PMID: 33214855 DOI: 10.21315/tlsr2020.31.3.4
    Simian betaretrovirus serotype-2 (SRV-2) is an important pathogenic agent in Asian macaques. It is a potential confounding variable in biomedical research. SRV-2 also provides a valuable viral model compared to other retroviruses which can be used for understanding many aspects of retroviral-host interactions and immunosuppression, infection mechanism, retroviral structure, antiretroviral and vaccine development. In this study, we isolated the gene encoding reverse transcriptase enzyme (RT) of SRV-2 that infected Indonesian cynomolgus monkey (Mf ET1006) and predicted the three dimensional structure model using the iterative threading assembly refinement (I-TASSER) computational programme. This SRV-2 RT Mf ET1006 consisted of 547 amino acids at nucleotide position 3284-4925 of whole genome SRV-2. The polymerase active site located in the finger/palm subdomain characterised by three conserved catalytic aspartates (Asp90, Asp165, Asp166), and has a highly conserved YMDD motif as Tyr163, Met164, Asp165 and Asp166. We estimated that this SRV-2 RT Mf ET1006 structure has the accuracy of template modelling score (TM-score 0.90 ± 0.06) and root mean square deviation (RMSD) 4.7 ± 3.1Å, indicating that this model can be trusted and the accuracy can be seen from the appearance of protein folding in tertiary structure. The superpositionings between SRV-2 RT Mf ET1006 and Human Immunodeficiency Virus-1 (HIV-1) RT were performed to predict the structural in details and to optimise the best fits for illustrations. This SRV-2 RT Mf ET1006 structure model has the highest homology to HIV-1 RT (2B6A.pdb) with estimated accuracy at TM-score 0.911, RMSD 1.85 Å, and coverage of 0.953. This preliminary study of SRV-2 RT Mf ET1006 structure modelling is intriguing and provide some information to explore the molecular characteristic and biochemical mechanism of this enzyme.
    Matched MeSH terms: Protein Folding
  19. NS3 protease of Langat tick-borne flavivirus cleaves serine protease substrates
    Scaramozzino N, Crance JM, Drouet C, Roebuck JP, Drouet E, Jouan A, et al.
    Biochem Biophys Res Commun, 2002 May 31;294(1):16-22.
    PMID: 12054734
    Langat (LGT) virus, initially isolated in 1956 from ticks in Malaysia, is a naturally occurring nonpathogenic virus with a very close antigenicity to the highly pathogenic tick-borne encephalitis (TBE) Western subtype virus and TBE Far Eastern subtype virus. NS3, the second largest viral protein of LGT virus, is highly conserved among flaviviruses and contains a characteristic protease moiety (NS3 pro). NS3 pro represents an attractive target for anti-protease molecules against TBE virus. We report herein a purification method specially designed for NS3 pro of LGT using a strategy for proper refolding coupled with the enzymatic characterisation of the protein. Different p-nitroanilide substrates, defined on canonic sequences for their susceptibility to Ser-protease, were applied to the proteolytic assays of the protein. The highest values were obtained from substrates containing an Arg or Lys (amino acid) residue at the P1 position. This purification method will facilitate the future development of reliable testing procedures for anti-proteases directed to NS3 proteins.
    Matched MeSH terms: Protein Folding
  20. Functional role of Ile264 in CYP2C8: mutations affect haem incorporation and catalytic activity
    Singh R, Ting JG, Pan Y, Teh LK, Ismail R, Ong CE
    Drug Metab. Pharmacokinet., 2008;23(3):165-74.
    PMID: 18574320
    The work described in this study aimed to express CYP2C8 wild-type and mutant proteins in bacterial expression system and to use the expressed proteins to investigate the structural and functional consequences of a reported allele CYP2C8(*)4 (carrying Ile264Met substitution) on protein activity. Ile264 was replaced by three different amino acids resulting in three mutant constructs, 2C8I264M, 2C8I264R and 2C8I264D. The presence of isoleucine at position 264 in CYP2C8 was found to be important for proper haem insertion and protein folding; whereas bulkier or charged residues were highly disruptive resulting in inactive proteins with minimum spectral and catalytic activities. This was evidenced from the low levels of Soret peak at 450 nm and negligible levels of tolbutamide methylhydroxylase activity. Kinetic study using paclitaxel indicated that all three mutants exhibited only 9.7 to 35.4% of the activity level observed in the wild-type. In addition, the mutants were more sensitive to proteinase K digestion, indicating a possible alteration of conformation. The combined effects of protein instability and compromised catalytic activity resulted in defective CYP2C8 protein which may have clinical implications in carriers of CYP2C8*4, particularly in terms of their capacity to clear potent drugs and their susceptibility to adverse drug reactions.
    Matched MeSH terms: Protein Folding
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