scholarly journals Computational-approach understanding the structure-function prophecy of Fibrinolytic Protease RFEA1 from Bacillus cereus RSA1

PeerJ ◽  
2021 ◽  
Vol 9 ◽  
pp. e11570
Author(s):  
Chhavi Sharma ◽  
Arti Nigam ◽  
Rajni Singh
Keyword(s):  
Structure Function ◽  
Bacillus Cereus ◽  
Structural Model ◽  
Three Dimensional ◽  
Conformational Space ◽  
Dimensional Structure ◽  
After Effects ◽  
Short Life Span ◽  
Therapeutic Enzymes

Microbial fibrinolytic proteases are therapeutic enzymes responsible to ameliorate thrombosis, a fatal cardiac-disorder which effectuates due to excessive fibrin accumulation in blood vessels. Inadequacies such as low fibrin specificity, lethal after-effects and short life-span of available fibrinolytic enzymes stimulates an intensive hunt for novel, efficient and safe substitutes. Therefore, we herewith suggest a novel and potent fibrinolytic enzyme RFEA1 from Bacillus cereus RSA1 (MK288105). Although, attributes such as in-vitro purification, characterization and thrombolytic potential of RFEA1 were successfully accomplished in our previous study. However, it is known that structure-function traits and mode of action significantly aid to commercialization of an enzyme. Also, predicting structural model of a protein from its amino acid sequence is challenging in computational biology owing to intricacy of energy functions and inspection of vast conformational space. Our present study thus reports In-silico structural-functional analysis of RFEA1. Sequence based modelling approaches such as—Iterative threading ASSEmbly Refinement (I-TASSER), SWISS-MODEL, RaptorX and Protein Homology/analogY Recognition Engine V 2.0 (Phyre2) were employed to model three-dimensional structure of RFEA1 and the modelled RFEA1 was validated by structural analysis and verification server (SAVES v6.0). The modelled crystal structure revealed the presence of high affinity Ca1 binding site, associated with hydrogen bonds at Asp147, Leu181, Ile185 and Val187residues. RFEA1 is structurally analogous to Subtilisin E from Bacillus subtilis 168. Molecular docking analysis using PATCH DOCK and FIRE DOCK servers was performed to understand the interaction of RFEA1 with substrate fibrin. Strong RFEA1-fibrin interaction was observed with high binding affinity (−21.36 kcal/mol), indicating significant fibrinolytic activity and specificity of enzyme RFEA1. Overall, the computational research suggests that RFEA1 is a subtilisin-like serine endopeptidase with proteolytic potential, involved in thrombus hydrolysis.

scholarly journals Nanofiber-Mâché Hollow Ball Mimicking the Three-Dimensional Structure of a Cyst

Polymers ◽  
2021 ◽  
Vol 13 (14) ◽  
pp. 2273
Author(s):  
Wan-Ying Huang ◽  
Norichika Hashimoto ◽  
Ryuhei Kitai ◽  
Shin-ichiro Suye ◽  
Satoshi Fujita
Keyword(s):  
Three Dimensional ◽  
Hollow Structure ◽  
Dimensional Structure ◽  
The Novel ◽  
Fibrous Scaffold ◽  
Hydrogel Beads ◽  
Inner Surface ◽  
Intracranial Epidermoid ◽  
Hollow Ball

The occasional malignant transformation of intracranial epidermoid cysts into squamous cell carcinomas remains poorly understood; the development of an in vitro cyst model is urgently needed. For this purpose, we designed a hollow nanofiber sphere, the “nanofiber-mâché ball.” This hollow structure was fabricated by electrospinning nanofiber onto alginate hydrogel beads followed by dissolving the beads. A ball with approximately 230 mm3 inner volume provided a fibrous geometry mimicking the topography of the extracellular matrix. Two ducts located on opposite sides provided a route to exchange nutrients and waste. This resulted in a concentration gradient that induced oriented migration, in which seeded cells adhered randomly to the inner surface, formed a highly oriented structure, and then secreted a dense web of collagen fibrils. Circumferentially aligned fibers on the internal interface between the duct and hollow ball inhibited cells from migrating out of the interior, similar to a fish bottle trap. This structure helped to form an adepithelial layer on the inner surface. The novel nanofiber-mâché technique, using a millimeter-sized hollow fibrous scaffold, is excellently suited to investigating cyst physiology.

scholarly journals Protein Folding: Search for Basic Physical Models

2003 ◽  
Vol 3 ◽  
pp. 623-635 ◽  
Author(s):  
Ivan Y. Torshin ◽  
Robert W. Harrison
Keyword(s):  
Protein Folding ◽  
Tertiary Structure ◽  
Three Dimensional ◽  
Physical Models ◽  
Dimensional Structure ◽  
Computational Techniques ◽  
Linear Sequence ◽  
Physical Forces

How a unique three-dimensional structure is rapidly formed from the linear sequence of a polypeptide is one of the important questions in contemporary science. Apart from biological context ofin vivoprotein folding (which has been studied only for a few proteins), the roles of the fundamental physical forces in thein vitrofolding remain largely unstudied. Despite a degree of success in using descriptions based on statistical and/or thermodynamic approaches, few of the current models explicitly include more basic physical forces (such as electrostatics and Van Der Waals forces). Moreover, the present-day models rarely take into account that the protein folding is, essentially, a rapid process that produces a highly specific architecture. This review considers several physical models that may provide more direct links between sequence and tertiary structure in terms of the physical forces. In particular, elaboration of such simple models is likely to produce extremely effective computational techniques with value for modern genomics.

scholarly journals Functional Analysis of the Mycobacterium tuberculosis FAD-Dependent Thymidylate Synthase, ThyX, Reveals New Amino Acid Residues Contributing to an Extended ThyX Motif

2008 ◽  
Vol 190 (6) ◽  
pp. 2056-2064 ◽  
Author(s):  
Jonathan E. Ulmer ◽  
Yap Boum ◽  
Christopher D. Thouvenel ◽  
Hannu Myllykallio ◽  
Carol Hopkins Sibley
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Thymidylate Synthase ◽  
Three Dimensional ◽  
Pyrimidine Ring ◽  
Dimensional Structure ◽  
Directed Mutagenesis ◽  
Catalytic Residue ◽  
Amino Acid Residues ◽  
Insight Into

ABSTRACT A novel FAD-dependent thymidylate synthase, ThyX, is present in a variety of eubacteria and archaea, including the mycobacteria. A short motif found in all thyX genes, RHRX7-8S, has been identified. The three-dimensional structure of the Mycobacterium tuberculosis ThyX enzyme has been solved. Building upon this information, we used directed mutagenesis to produce 67 mutants of the M. tuberculosis thyX gene. Each enzyme was assayed to determine its ability to complement the defect in thymidine biosynthesis in a ΔthyA strain of Escherichia coli. Enzymes from selected strains were then tested in vitro for their ability to catalyze the oxidation of NADPH and the release of a proton from position 5 of the pyrimidine ring of dUMP. The results defined an extended motif of amino acids essential to enzyme activity in M. tuberculosis (Y44X24 H69X25R95HRX7 S105XRYX90R199 [with the underlined histidine acting as the catalytic residue and the underlined serine as the nucleophile]) and provided insight into the ThyX reaction mechanism. ThyX is found in a variety of bacterial pathogens but is absent in humans, which depend upon an unrelated thymidylate synthase, ThyA. Therefore, ThyX is a potential target for development of antibacterial drugs.

scholarly journals In Vitro Selection and Characterization of Hepatitis C Virus Serine Protease Variants Resistant to an Active-Site Peptide Inhibitor

2003 ◽  
Vol 77 (6) ◽  
pp. 3669-3679 ◽  
Author(s):  
Caterina Trozzi ◽  
Linda Bartholomew ◽  
Alessandra Ceccacci ◽  
Gabriella Biasiol ◽  
Laura Pacini ◽  
...  
Keyword(s):  
Hepatitis C Virus ◽  
Hepatitis C ◽  
Serine Protease ◽  
In Vitro Selection ◽  
Three Dimensional ◽  
Host Cells ◽  
Dimensional Structure ◽  
In Vitro Systems

ABSTRACT The hepatitis C virus (HCV) serine protease is necessary for viral replication and represents a valid target for developing new therapies for HCV infection. Potent and selective inhibitors of this enzyme have been identified and shown to inhibit HCV replication in tissue culture. The optimization of these inhibitors for clinical development would greatly benefit from in vitro systems for the identification and the study of resistant variants. We report the use HCV subgenomic replicons to isolate and characterize mutants resistant to a protease inhibitor. Taking advantage of the replicons' ability to transduce resistance to neomycin, we selected replicons with decreased sensitivity to the inhibitor by culturing the host cells in the presence of the inhibitor and neomycin. The selected replicons replicated to the same extent as those in parental cells. Sequence analysis followed by transfection of replicons containing isolated mutations revealed that resistance was mediated by amino acid substitutions in the protease. These results were confirmed by in vitro experiments with mutant enzymes and by modeling the inhibitor in the three-dimensional structure of the protease.

scholarly journals Secondary Structural Model of Human MALAT1 Reveals Multiple Structure–Function Relationships

2019 ◽  
Vol 20 (22) ◽  
pp. 5610 ◽  
Author(s):  
Phillip J. McCown ◽  
Matthew C. Wang ◽  
Luc Jaeger ◽  
Jessica A. Brown
Keyword(s):  
Structure Function ◽  
Noncoding Rna ◽  
Structural Model ◽  
Dynamic Structure ◽  
Nucleotide Polymorphisms ◽  
Rna Structures ◽  
Rna Modifications ◽  
Functional Roles ◽  
Conserved Core

Human metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) is an abundant nuclear-localized long noncoding RNA (lncRNA) that has significant roles in cancer. While the interacting partners and evolutionary sequence conservation of MALAT1 have been examined, much of the structure of MALAT1 is unknown. Here, we propose a hypothetical secondary structural model for 8425 nucleotides of human MALAT1 using three experimental datasets that probed RNA structures in vitro and in various human cell lines. Our model indicates that approximately half of human MALAT1 is structured, forming 194 helices, 13 pseudoknots, five structured tetraloops, nine structured internal loops, and 13 intramolecular long-range interactions that give rise to several multiway junctions. Evolutionary conservation and covariation analyses support 153 of 194 helices in 51 mammalian MALAT1 homologs and 42 of 194 helices in 53 vertebrate MALAT1 homologs, thereby identifying an evolutionarily conserved core that likely has important functional roles in mammals and vertebrates. Data mining revealed that RNA modifications, somatic cancer-associated mutations, and single-nucleotide polymorphisms may induce structural rearrangements that sequester or expose binding sites for several cancer-associated microRNAs. Our findings reveal new mechanistic leads into the roles of MALAT1 by identifying several intriguing structure–function relationships in which the dynamic structure of MALAT1 underlies its biological functions.

The Integrative Establishment of a Three-Dimensional Structure Model of Changling Gas Field in Changling Fault Depression

2014 ◽  
Vol 556-562 ◽  
pp. 3779-3782
Author(s):  
Xiao Yu Yu ◽  
Xue Li ◽  
Xiao Song Li ◽  
Guo Yi Zhang
Keyword(s):  
Structural Model ◽  
High Reliability ◽  
Three Dimensional ◽  
Physical Property ◽  
Modeling Technique ◽  
Dimensional Structure ◽  
Gas Field ◽  
3D Geological Modeling ◽  
Micro Features ◽  
Reservoir Description

The three-dimensional (3D) geological modeling technique which is considered as an important skill of fine reservoir description has been gaining more and more attention. On one hand, it can efficiently promote the transformation of reservoir description from two-dimensional (2D) to 3D, and from qualification to quantification as well. The 3D reservoir geological model can be used as basic geological knowledge in terms of adjusting well patterns and indicating remaining oil distribution, through reflecting the spatial distribution characteristics and the variation of the reservoir physical property. On the other hand, the 3D modeling technique specializes in the representation of local micro features in comparison of regular ways. This article aims at subtly describing the structural modeling of Changling gas field of Changling fault depression. The result of this case study shows that the establishment of structural model is consistent with the understanding of fault development which was proved during the process of producing gas, thus the structural model has high reliability. Therefore, the structural model is of great guiding significance for the design of new well and the well patter optimization.

Rational humanization of the powerful antithrombotic anti-GPIbα antibody: 6B4

2006 ◽  
Vol 96 (11) ◽  
pp. 671-684 ◽  
Author(s):  
Alexandre Fontayne ◽  
Karen Vanhoorelbeke ◽  
Inge Pareyn ◽  
Isabel Van Rompaey ◽  
Muriel Meiring ◽  
...  
Keyword(s):  
Ex Vivo ◽  
Three Dimensional ◽  
Vector System ◽  
Dimensional Structure ◽  
Fab Fragment ◽  
Variable Regions ◽  
Significant Prolongation ◽  
Von Willebrand ◽  
First Time

SummaryFab-fragments of the monoclonal antibody 6B4, raised against human glycoprotein Ibα (GPIbα), have a powerful antithrombotic effect in baboons by blocking the GPIbα binding site for von Willebrand factor (VWF), without significant prolongation of the skin bleeding time. In order to bring this antibody to the clinic,we here humanized for the first time an anti-human GPIbα by variable-domain resurfacing guided by computer modeling. First, the genes coding for the variable regions of the heavy and light chains of 6B4 were cloned and sequenced. Based on this,a three-dimensional structure of the Fv-fragment was constructed by using homology-based modeling, and with this and comparison with antibodies with known structure,”murine” putative immunogenic residues which are exposed, were changed for “human-like” residues. The humanized Fab-fragment, h6B4-Fab, was constructed in the pKaneo vector system, expressed and purified and showed in vitro an unaltered, even slightly higher binding affinity for its antigen than the murine form as determined by different ELISA set-ups and surface plasmon resonance. Finally, injection of doses of 0.1 to 1.5 mg/kg of h6B4-Fab in baboons showed that both pharmacokinetics and ex-vivo bio-activity of the molecule were to a large extent preserved.In conclusion, the method used here to humanize 6B4 by resurfacing resulted in a fully active derivative, which is now ready for further development.

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