Arrangement Of Amino Acids Research Articles

Preparation and evaluation a mixed-mode stationary phase with C18 and 2-methylindole for HPLC.

A modified C18 column (Silpr-2MI-C18) was prepared using 2-methylindole and C18 reagent. The extent of C18 hydrocarbon chain, conjugative rings and anion exchange site provided multiple retention mechanisms, including reversed-phase liquid chromatography (RPLC), π-π interaction, hydrophilic interaction liquid chromatography (HILIC) and anion exchange chromatography (AEC). The separation of protected amino acids was investigated on the commercial C18 and Silpr-2MI-C18 columns, while the chromatographic conditions, including methanol content and pH of the mobile phase, were studied. The separation arrangement of the hydrophilic amino acids was different on the Silpr-2MI-C18 column compared to the commercial C18 column under RPLC mode. Furthermore, these amino acids were separated on the Silpr-2MI-C18 column under HILIC mode. The modified C18 column was employed to separate amino acids, alkylbenzenes and polycyclic aromatic hydrocarbons under RPLC mode and inorganic anion under AEC mode. The results confirm that this new stationary phase of RPLC/HILIC/AEC has multiple interactions with different analytes. Effective retention of biological samples was found on the Silpr-2MI-C18 column by comparing the results obtained from the commercial C18 column.

Phylogenetic analysis of HPAI H5N1 virus from duck swab specimens in Indonesia.

Objective:A phylogenetic study was carried out on the avian influenza virus (AIV) isolated from a disease outbreak in Sidenreng Rappang Regency, South Sulawesi, Indonesia, in 2018.Material and Methods:Oropharyngeal swabs and organ samples were obtained from ducks that showed clinical symptoms: torticollis, fascial edema, neurological disorders, the corneas appear cloudy, and death occurs less than 1 day after symptoms appear. In this study, isolate A/duck/Sidenreng Rappang/07180110-11/2018 from duck was sequenced and characterized.Results:It was found that each gene segment of the virus has the highest nucleotide homology to the Indonesian highly pathogenic avian influenza (HPAI) H5N1 clade 2.3.2.1c. Multiple alignments of the sample Hemagglutinin (HA) gene with the avian influenza references virus showed that the pattern of amino acid arrangement in the cleavage site PQRERRRK-RGLF is the characteristic of the HPAI virus. In addition, the HA gene contained Q222 (glutamine) and G224 (glycine), signifying a high affinity to avian receptor binding specificity (SA α2,3 Gal). Furthermore, there was no genetic reassortment of this virus based on the phylogenetic analysis of HA, NA, PB1, PB2, PA, NP, M, and NS genes.Conclusion:The HPAI H5N1 clade 2.3.2.1c virus was identified in duck farms in South Sulawesi, Indonesia.

Pemodelan Homologi Komparatif Struktur 3d Protein dalam Desain dan Pengembangan Obat

One of the strategies applied in the initial steps of drug design and discovery is by utilizing the availability of 3D protein structures. However, most of functional protein structures have not been fully obtained experimentally to date. This is due to the difficulty of procedures, the high cost and the length of time required from structural biology experiments such as X-ray crystallography. Comparative homology modeling is a computational method that is proven to be accurate in predicting the structure of 3D target proteins with a ratio of 30% similarity to the arrangement of amino acids in the structure of the template proteins. This review attempts to explain comparative homology modeling methods such as MODELLER, PHYRE2, SWISS-MODEL, its iterative steps in predicting and building 3D protein target models, its evaluation and validation of protein models as well as the examples of their application in drug design and development targeting GPCR proteins, Zika virus RNA polymerase and HIV protease. With this, comparative homology modeling can be very useful to accelerate research on drug design and development in dealing with diseases and health problems that still exist in the society.

Design of new truncated derivatives based on direct and reverse mirror repeats of first six residues of Caerin 4 antimicrobial peptide and evaluation of their activity and cytotoxicity.

Caerin 4 is a family of AMPs isolated from the frog called Litoria caerulea. In silico drug designing methods and using machine learning algorithms for AMPs design can reduce their usage restrictions such as production costs and the time required for investigation of their activity and toxicity. In this study, two short peptides were designed based on direct and reverse mirror repeats of GLWQKI conserved sequence from Caerin 4 family that called dCar12 and rCar12. Also, Caerin 4.1 was synthesized without primary GLWQKI sequence and named Car7-23 . Following the synthesis of peptides, their antimicrobial properties, cytotoxicity, secondary structure, and mode of action were further evaluated. Results indicated that rCar12 had a good antibacterial activity (at an MIC of 3.9-62.5µg/ml), while Car7-23 did not have any antimicrobial properties. Cytotoxicity of rCar12 at MICs range was <5%, which is much less than Caerin 4.1. In conclusion, rCar12 with reverse mirror repeat has different functional properties compared with dCar12. These results corroborate the fact that in two peptides with identical residues and length, the position and arrangement of amino acids are very important concerning peptide function. Moreover, GLWQKI sequence is highly crucial for the antimicrobial activity of Caerin 4 antimicrobial peptide family.

The Nature and Arrangement of Pentatricopeptide Domains and the Linker Sequences Between Them

The tricopeptide (amino acid number in the 30s) repeats constitute some of the most common amino acid repeats in proteins of diverse organisms. The most important representatives of this class are the 34-residue and 35-residue repeats, eponymously known as tetratricopeptide repeat (TPR) and pentatricopeptide repeat (PPR), respectively. The unit motif of both consists of a pair of alpha helices. As members of the large, all-helical repeat classes, TPR and PPR share structural similarities, but also play specific roles in protein function. In this study, a comprehensive bioinformatic analysis of the PPR units and the linkers that connect them was conducted. The results suggested the existence of PPR repeats of various formats, as well as smaller, PPR-unrelated repeats. Besides their length, these repeats differed in amino acid arrangements and location of key amino acids. These findings provide a broader and unified perspective of the pentatricopeptide family while raising provocative questions about the assembly and evolution of these domains.

Helix-helix interaction viewed in an angle frame indicates a role of the size of sidechains in packing

Three-dimensional packing is essential for protein folding. Homology alignment and de nova methods have been used to develop algorithms that would predict protein packing in proteins. Even though hydrophobic cores of proteins are formed through the packing of side chains of hydrophobic residues, the stereological arrangement of individual amino acids and in three dimensional hydrophobic cores is rather difficult to determine. In order to simplify the description of a protein core packing, in this manuscript, we have determined the relationship between angles, distances, and residue usage between two helices. Data from this analysis demonstrate that the contact residues determine the angle and the distance between two helical elements. This approach provides a means to predict the three-dimensional packing of helices in proteins and allows for an understanding of the biological interaction within proteins and among proteins based on surface contact residue parameters.

Induced Perturbation Network and tiling for modeling the L55P Transthyretin amyloid fiber

Abstract Protein structures are complex spatial systems, formed by the three-dimensional arrangement of amino acids, that interact through atomic contacts. It is fundamental to understand the perturbation mechanisms associated with the amino acid mutations that lead to changes in a protein dynamics, as such mutations can lead to diseases. We present a methodology based on Amino Acid Networks and the use of Induced Perturbation Networks to infer the impact of amino acid mutations on a protein’s dynamics and the use of a tiling formalism to model protein aggregation. We apply this methodology to the case study of the L55P Transthyretin (TTR) variant, one of the most pathogenic mutations of TTR, due to its increased tendency to aggregate into amyloid fibers. We show that another pathogenic variant, V30M TTR, produces different results, reflecting a different pathogenic mechanism compared to L55P TTR and that the results differ for pathogenic and non-pathogenic variants (L55P and V30M TTR pathogenic variants versus Tl 19Y and Tl 19M non-pathogenic variants).

Comparison of partial and full &lt;em&gt;VP2&lt;/em&gt; gene sequences of parvovirus from domestic cats in Turkey

Parvoviruses are ubiquitous pathogens that cause a fatal disease in cats and are able to mutate for cross-species transmission. Both the feline panleukopenia virus (FPV) and the canine parvovirus (CPV), with their antigenic variants, induce a disease in cats that presents with similar signs. The aim of this study was to determine the presence of parvoviruses in blood and exudate samples from five clinically symptomatic cats (from Ankara, Turkey). The gene coding for the VP2 structural capsid protein of the obtained parvoviruses was amplified by polymerase chain reaction (PCR), purified and partially or nearly full-length sequenced. The maximum likelihood (ML) method was used for molecular characterization. Phylogenetic analysis based on nearly full-length sequencing of the VP2 gene and amino acid arrangement showed that four of the viral strains were closely related and localized in the same FPV cluster. The fifth strain found was located in the same cluster but on a separate branch. Viral field strains were included in the CPV-2 group as determined by partial genome analysis: four fitted in the CPV-2c, and one in a separate clade within the CPV-2b group. To our knowledge, this is the first report that details nearly full-length VP2 gene characterisation in Turkish cats. Overall, nearly full-length VP2 contrasts were more effective to determine the origin of parvovirus strains, than partial length comparisons.

Tuning the Structure and Chiroptical Properties of Gold Nanoparticle Single Helices via Peptide Sequence Variation.

Just as peptide function is determined by the position, sequence, and overall arrangement of constituent amino acids, the optical properties of nanoparticle (NP) assemblies are influenced by the size, dimensions, and arrangement of constituent NPs. In this work, we demonstrate that peptide sequence can be programmed to direct the structure and chiroptical activity of chiral helical gold NP (AuNP)superstructures, a growing class of chiral nanomaterials with potential in sensing, detection, and optics-based applications. Gold-binding peptide conjugate families, C18-(PEPAuM,x)2 and C18-(PEPAuM-ox,x)2, that differ in the position (x = 7, 9, and 11) of methionine (M)/methionine sulfoxide (M-ox) within the peptide sequences (PEPAu = AYSSGAPPMPPF/PEPAuM-ox = AYSSGAPPMoxPPF) are employed to control the aspect ratio and size of AuNPs within helical NP assemblies. Computational modeling reveals that the amino acid variations have a profound effect on peptide-AuNP interactions that ultimately lead to control over NP size. C18-(PEPAuM,x)2 (x = 7, 9, and 11) yield irregular double-helical superstructures comprising spherical AuNPs, while C18-(PEPAuM-ox,x)2 (x = 9, 11) yield single-helical assemblies comprising oblong or rod-shaped AuNPs. Further, component AuNPs are larger when M/M-ox is placed at x = 11, while smaller component AuNPs are observed when M/M-ox is placed at x = 7. Changes in nanoscale structures manifest themselves in observable differences in chiroptical signal intensity. Ultimately, we achieve dramatic variance in the structure and properties of chiral AuNP superstructures via simple molecular-level tuning of peptide primary sequence.

Presence of a basic secretory protein in xylem sap and shoots of poplar in winter and its physicochemical activities against winter environmental conditions.

XSP25, previously shown to be the most abundant hydrophilic protein in xylem sap of Populus nigra in winter, belongs to a secretory protein family in which the arrangement of basic and acidic amino acids is conserved between dicotyledonous and monocotyledonous species. Its gene expression was observed at the same level in roots and shoots under long-day conditions, but highly induced under short-day conditions and at low temperatures in roots, especially in endodermis and xylem parenchyma in the root hair region of Populus trichocarpa, and its protein level was high in dormant buds, but not in roots or branches. Addition of recombinant PtXSP25 protein mitigated the denaturation of lactate dehydrogenase by drying, but showed only a slight effect on that caused by freeze-thaw cycling. Recombinant PtXSP25 protein also showed ice recrystallization inhibition activity to reduce the size of ice crystals, but had no antifreezing activity. We suggest that PtXSP25 protein produced in shoots and/or in roots under short-day conditions and at non-freezing low temperatures followed by translocation via xylem sap to shoot apoplast may protect the integrity of the plasma membrane and cell wall functions from freezing and drying damage in winter environmental conditions.

Mutant power resolves protein shapes

Clever calculations on mutated proteins reveal spatial arrangement of amino acids.

Drug ReposER: a web server for predicting similar amino acid arrangements to known drug binding interfaces for potential drug repositioning.

A common drug repositioning strategy is the re-application of an existing drug to address alternative targets. A crucial aspect to enable such repurposing is that the drug's binding site on the original target is similar to that on the alternative target. Based on the assumption that proteins with similar binding sites may bind to similar drugs, the 3D substructure similarity data can be used to identify similar sites in other proteins that are not known targets. The Drug ReposER (DRug REPOSitioning Exploration Resource) web server is designed to identify potential targets for drug repurposing based on sub-structural similarity to the binding interfaces of known drug binding sites. The application has pre-computed amino acid arrangements from protein structures in the Protein Data Bank that are similar to the 3D arrangements of known drug binding sites thus allowing users to explore them as alternative targets. Users can annotate new structures for sites that are similarly arranged to the residues found in known drug binding interfaces. The search results are presented as mappings of matched sidechain superpositions. The results of the searches can be visualized using an integrated NGL viewer. The Drug ReposER server has no access restrictions and is available at http://mfrlab.org/drugreposer/.

Ultrafast Dynamics of Heliorhodopsins.

Heliorhodopsins (HeR) constitute a new rhodopsin family and show only <15% sequence identities with type-1 and type-2 rhodopsins. The large difference in amino acid sequence between HeRs and other rhodopsins raises a question whether their biological function is triggered by efficient and rapid photoisomerization of the retinal chromophore as in the case of other rhodopsins. We performed femtosecond time-resolved absorption measurements of two HeRs, HeR 48C12 and HeR from Thermoplasmatales archaeon SG8-52-1. Both HeRs exhibit excited-state absorption around 480 nm and stimulated emission in the >650 nm region, and these transient signals decay concomitantly with appearance of photoproduct absorption on a subpicosecond time scale. The observed spectral change indicates that ultrafast retinal photoisomerization proceeds in the femtosecond time region. The transient spectra and dynamics of HeRs are surprisingly similar to those of type-1 rhodopsins, despite remarkable differences in amino acid arrangement in the hydrophobic region of the retinal binding site.

Antimicrobial Peptides with High Proteolytic Resistance for Combating Gram-Negative Bacteria.

Poor proteolytic resistance is an urgent problem to be solved in the clinical application of antimicrobial peptides (AMPs), yet common solutions, such as complicated chemical modifications and utilization of d-amino acids, greatly increase the difficulty and cost of producing AMPs. In this work, a set of novel peptides was synthesized based on an antitrypsin/antichymotrypsin hydrolytic peptide structure unit (XYPX) n (X represents I, L, and V; Y represents R and K), which was designed using a systematic natural amino acid arrangement. Of these peptides, 16 with seven repeat units had the highest average selectivity index (GMSI = 99.07) for all of the Gram-negative bacteria tested and remained highly effective in combating Escherichia coli infection in vivo. Importantly, 16 also had dramatic resistance to a high concentration of trypsin/chymotrypsin hydrolysis and exerted bactericidal activity through a membrane-disruptive mechanism. Overall, these findings provide new approaches for the development of antiprotease hydrolytic peptides that target Gram-negative bacteria.

Identification of G-Quadruplex-Binding Protein from the Exploration of RGG Motif/G-Quadruplex Interactions.

The arginine/glycine-rich region termed the RGG domain is usually found in G-quadruplex (G4)-binding proteins and is important in G4-protein interactions. Studies on the binding mechanism of RGG domains found that small segments (RGG motif) inside the domain contribute greatly to the G4 binding affinity. However, unlike the entire RGG domains that have been broadly explored, the role of the RGG motif remains obscure, with very limited study. Herein, to clarify the role of the RGG motif in G4-protein interactions, we systematically investigated the binding affinity and mode between RGG-motif peptides and G4s. The internal arrangement of RGG repeats and gap amino acids played a more crucial role in the G4-binding mechanism than a critical number of RGG repeats. Arginines and phenylalanines at the exact position of the RGG motif might enable additional hydrogen bonding and π-stacking interaction with nucleobases and strengthen the binding of G4. Impressively, proceeding from a G4-binding RGG peptide, 12, discovered above, we identified the cold-inducible RNA-binding protein (CIRBP) as a new G4 DNA-binding protein both in vitro and in cells. In addition, we found that the key amino acids for G4 binding in peptide 12 and CIRBP were highly similar, and peptide 12 clearly played a key role in the G4 binding of CIRBP. This report is the first in which a G4-binding protein was identified from exploration of the G4-binding RGG motif. Our findings suggest a novel strategy for discovering new G4-binding proteins by exploring key peptide segments.

Radical scavenging property of a novel peptide derived from C-terminal SOD domain of superoxide dismutase enzyme in Arthrospira platensis

Superoxide dismutase (SOD) is an evolutionary conserved detoxification enzyme and powerful antioxidant which defends against the elevated ROS that are induced by various stresses. Arthrospira platensis (Ap) is known for its antioxidant-mediated immunostimulant properties, but there is no report on the SOD dependent antioxidant mechanism. Therefore, in this study, we have analysed the effect of H2O2 on growth and pigment composition in spirulina. Results showed that spirulina exposed to 10 mM H2O2 showed elevated growth pattern as well as increase in chlorophyll pigment composition especially during early days of exposure. Gene expression results showed that the expression profile of ApSOD during oxidative stress stimulated by 10 mM H2O2 at different time intervals (0, 5, 10, 15 and 20 days) with highest expression on day 10 post-exposure. Together, the results confirmed the antioxidant role of ApSOD in spirulina during oxidative stress induced by H2O2. Based on the amino acid arrangement and composition, we have predicted a short peptide 160LGLDVWEHAYYL171 (LL12) from the catalytic centre of C-terminal SOD domain; further the peptide was synthesized. Antioxidant assays showed that LL12 peptide critically involved in radical scavenging mechanism. Also, LL12 peptide reduced the intracellular ROS level in H2O2 exposed leucocytes at a concentration of 12.5 μM. Cytotoxicity assay was performed on human leucocytes which showed that LL12 did not exhibit any cytotoxic activity against any of the leucocytes population. Overall, the study highlights the radical scavenging property of a novel short peptide derived from the C-terminal domain of ApSOD which have the potential to develop as a biopharmaceutical drug.

A novel improved prediction of protein structural class using deep recurrent neural network

For last few decades, sequence arrangement of amino acids have been utilized for the prediction of protein secondary structure. Recent methods have applied high dimensional natural language based features in machine learning models. Performance measures of machine learning based models are significantly affected by data size and data dimensionality. It is a huge challenge to develop a generic model which can be trained to perform both for small and large sized datasets in a low dimensional framework. In the present research, we suggest a low dimensional representation for both small and large sized datasets. A hybrid space of Atchley’s factors II, IV, V, electron ion interaction potential and SkipGram based word2vec have been employed for amino acid sequence representation. Subsequently Stockwell transformation is applied to the representation to preserve features both in time and frequency domains. Finally, deep gated recurrent network with dropout, categorical-cross entropy error estimation and Adam optimization is used for classification purpose. The introduced method results in better prediction accuracies for both small (204,277, and 498) and large sized (PDB25, Protein 640 and FC699) bench mark data sets of low sequence similarity (25–40%). The obtained classification accuracies for PDB25, 640, FC699, 498, 277, 204 datasets are 84.2%, 94.31%, 93.1%, 95.9%, 94.5% and 85.36% respectively. The major contributions in this research is that, for the first time, we verify the protein secondary structural class prediction in a very low dimensional (18-D) feature space with a novel feature representation method. Secondly, we also verify for the first time, the behaviour of deep networks for low dimensional small sized data sets.

Role of solvent accessibility for aggregation-prone patches in protein folding

The arrangement of amino acids in a protein sequence encodes its native folding. However, the same arrangement in aggregation-prone regions may cause misfolding as a result of local environmental stress. Under normal physiological conditions, such regions congregate in the protein’s interior to avoid aggregation and attain the native fold. We have used solvent accessibility of aggregation patches (SAAPp) to determine the packing of aggregation-prone residues. Our results showed that SAAPp has low values for native crystal structures, consistent with protein folding as a mechanism to minimize the solvent accessibility of aggregation-prone residues. SAAPp also shows an average correlation of 0.76 with the global distance test (GDT) score on CASP12 template-based protein models. Using SAAPp scores and five structural features, a random forest machine learning quality assessment tool, SAAP-QA, showed 2.32 average GDT loss between best model predicted and actual best based on GDT score on independent CASP test data, with the ability to discriminate native-like folds having an AUC of 0.94. Overall, the Pearson correlation coefficient (PCC) between true and predicted GDT scores on independent CASP data was 0.86 while on the external CAMEO dataset, comprising high quality protein structures, PCC and average GDT loss were 0.71 and 4.46 respectively. SAAP-QA can be used to detect the quality of models and iteratively improve them to native or near-native structures.

RNA-Seq in the discovery of a sparsely expressed scent-determining monoterpene synthase in lavender (Lavandula).

Using RNA-Seq, we cloned and characterized a unique monoterpene synthase responsible for the formation of a scent-determining S-linalool constituent of lavender oils from Lavandula × intermedia. Several species of Lavandula produce essential oils (EOs) consisting mainly of monoterpenes including linalool, one of the most abundant and scent-determining oil constituents. Although R-linalool dominates the EOs of lavenders, varying amounts (depending on the species) of the S-linalool enantiomer can also be found in these plants. Despite its relatively low abundance, S-linalool contributes a sweet, pleasant scent and is an important constituent of lavender EOs. While several terpene synthase genes including R-linalool synthase have been cloned from lavenders many important terpene synthases including S-linalool synthase have not been described from these plants. In this study, we employed RNA-Seq and other complementary sequencing data to clone and functionally characterize the sparsely expressed S-linalool synthase cDNA (LiS-LINS) from Lavandula × intermedia. Recombinant LiS-LINS catalyzed the conversion of the universal monoterpene precursor geranyl diphosphate to S-linalool as the sole product. Intriguingly, LiS-LINS exhibited very low (~ 30%) sequence similarity to other Lavandula terpene synthases, including R-linalool synthase. However, the predicted 3D structure of this protein, including the composition and arrangement of amino acids at the active site, is highly homologous to known terpene synthase proteins. LiS-LINS transcripts were detected in flowers, but were much less abundant than those corresponding to LiR-LINS, paralleling enantiomeric composition of linalool in L. × intermedia oils. These data indicate that production of S-linalool is at least partially controlled at the level of transcription from LiS-LINS. The cloned LiS-LINS cDNA may be used to enhance oil composition in lavenders and other plants through metabolic engineering.

Protein-Cross-Linking zur Aufkl\xe4rung von komplexen Strukturen

Cryo-electron microscopy (cryo-EM) can solve structures of highly dynamic macromolecular complexes. To characterize less well defined regions in cryo-EM images, cross-linking coupled with mass spectrometry (CX-MS) provides valuable information on the arrangement of domains and amino acids. CX-MS involves covalent linkage of protein residues close to each other and identifying these connections by mass spectrometry. Here, we summarise the advances of CX-MS and its integration with cryo-EM for structural reconstruction.