scholarly journals Mechanism of nucleotide discrimination by the translesion synthesis polymerase Rev1

2021 ◽  
Author(s):  
Tyler M Weaver ◽  
Timothy H Click ◽  
Thu H Khoang ◽  
Todd M Washington ◽  
Pratul Agarwal ◽  
...  
Keyword(s):  
Hydrogen Bonds ◽  
Dna Synthesis ◽  
Active Site ◽  
Side Chain ◽  
Translesion Dna Synthesis ◽  
X Ray Crystallography ◽  
Abasic Sites ◽  
Dna Base ◽  
Dynamics Simulations ◽  
Insight Into

Rev1 is a translesion DNA synthesis (TLS) polymerase involved in the bypass of adducted-guanine bases and abasic sites during DNA replication. During damage bypass, Rev1 utilizes a protein-template mechanism of DNA synthesis, where the templating DNA base is evicted from the Rev1 active site and replaced by an arginine side chain that preferentially binds incoming dCTP. Here, we utilize X-ray crystallography and molecular dynamics simulations to obtain structural insight into the dCTP specificity of Rev1. We show the Rev1 R324 protein-template forms sub-optimal hydrogen bonds with incoming dTTP, dGTP, and dATP that prevents Rev1 from adopting a catalytically competent conformation. Additionally, we show the Rev1 R324 protein-template forms optimal hydrogen bonds with incoming rCTP. However, the incoming rCTP adopts an altered sugar pucker, which prevents the formation of a catalytically competent Rev1 active site. This work provides novel insight into the mechanisms for nucleotide discrimination by the TLS polymerase Rev1.

scholarly journals Crystal Structure of Escherichia coli Agmatinase: Catalytic Mechanism and Residues Relevant for Substrate Specificity

2021 ◽  
Vol 22 (9) ◽  
pp. 4769
Author(s):  
Pablo Maturana ◽  
María S. Orellana ◽  
Sixto M. Herrera ◽  
Ignacio Martínez ◽  
Maximiliano Figueroa ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Active Site ◽  
Catalytic Mechanism ◽  
Conformational Dynamics ◽  
High Specificity ◽  
Arginine Decarboxylase ◽  
Space Groups ◽  
X Ray Crystallography ◽  
High Dynamics ◽  
Dynamics Simulations

Agmatine is the product of the decarboxylation of L-arginine by the enzyme arginine decarboxylase. This amine has been attributed to neurotransmitter functions, anticonvulsant, anti-neurotoxic, and antidepressant in mammals and is a potential therapeutic agent for diseases such as Alzheimer’s, Parkinson’s, and cancer. Agmatinase enzyme hydrolyze agmatine into urea and putrescine, which belong to one of the pathways producing polyamines, essential for cell proliferation. Agmatinase from Escherichia coli (EcAGM) has been widely studied and kinetically characterized, described as highly specific for agmatine. In this study, we analyze the amino acids involved in the high specificity of EcAGM, performing a series of mutations in two loops critical to the active-site entrance. Two structures in different space groups were solved by X-ray crystallography, one at low resolution (3.2 Å), including a guanidine group; and other at high resolution (1.8 Å) which presents urea and agmatine in the active site. These structures made it possible to understand the interface interactions between subunits that allow the hexameric state and postulate a catalytic mechanism according to the Mn2+ and urea/guanidine binding site. Molecular dynamics simulations evaluated the conformational dynamics of EcAGM and residues participating in non-binding interactions. Simulations showed the high dynamics of loops of the active site entrance and evidenced the relevance of Trp68, located in the adjacent subunit, to stabilize the amino group of agmatine by cation-pi interaction. These results allow to have a structural view of the best-kinetic characterized agmatinase in literature up to now.

WHICH IS THE PROTON-SHUTTLE IN ISOXANTHOPTERIN DEAMINASE? QM/MM MD UNDERSTANDING

2013 ◽  
Vol 12 (08) ◽  
pp. 1341002 ◽  
Author(s):  
XIN ZHANG ◽  
MING LEI
Keyword(s):  
Crystal Structure ◽  
Molecular Dynamics ◽  
Hydrogen Bonds ◽  
Proton Transfer ◽  
Glutamic Acid ◽  
Active Site ◽  
Nucleophilic Attack ◽  
Zinc Ions ◽  
Initial Model ◽  
Dynamics Simulations

The deamination process of isoxanthopterin catalyzed by isoxanthopterin deaminase was determined using the combined QM(PM3)/MM molecular dynamics simulations. In this paper, the updated PM3 parameters were employed for zinc ions and the initial model was built up based on the crystal structure. Proton transfer and following steps have been investigated in two paths: Asp336 and His285 serve as the proton shuttle, respectively. Our simulations showed that His285 is more effective than Aap336 in proton transfer for deamination of isoxanthopterin. As hydrogen bonds between the substrate and surrounding residues play a key role in nucleophilic attack, we suggested mutating Thr195 to glutamic acid, which could enhance the hydrogen bonds and help isoxanthopterin get close to the active site. The simulations which change the substrate to pterin 6-carboxylate also performed for comparison. Our results provide reference for understanding of the mechanism of deaminase and for enhancing the deamination rate of isoxanthopterin deaminase.

scholarly journals Molecular basis of flexible peptide recognition by an antibody

2020 ◽  
Vol 167 (4) ◽  
pp. 343-345
Author(s):  
Koki Makabe
Keyword(s):  
Hydrogen Bonds ◽  
High Specificity ◽  
Recognition Mechanism ◽  
Intramolecular Hydrogen Bonds ◽  
Peptide Recognition ◽  
X Ray Crystallography ◽  
Biophysical Techniques ◽  
Antibody Design ◽  
Dynamics Simulations ◽  
Intramolecular Hydrogen

Abstract Antibodies can recognize various types of antigens with high specificity and affinity and peptide is one of their major targets. Understanding an antibody’s molecular recognition mechanism for peptide is important for developing clones with a higher specificity and affinity. Here, the author reviews recent progresses in flexible peptide recognition by an antibody using several biophysical techniques, including X-ray crystallography, molecular dynamics simulations and calorimetric measurements. A set of two reports highlight the importance of intramolecular hydrogen bonds that form in an unbound flexible state. Such intramolecular hydrogen bonds restrict the fluctuation of the peptide and reduce the conformational entropy, resulting in the destabilization of the unbound state and increasing the binding affinity by increasing the free energy change. These detailed analyses will aid in the antibody design in the future.

scholarly journals Molecular Dynamics Insight into the Lipid II Recognition by Type A Lantibiotics: Nisin, Epidermin, and Gallidermin

Micromachines ◽  
2021 ◽  
Vol 12 (10) ◽  
pp. 1169
Author(s):  
Irina Panina ◽  
Amir Taldaev ◽  
Roman Efremov ◽  
Anton Chugunov
Keyword(s):  
Molecular Dynamics ◽  
Side Chain ◽  
Amino Acid Residues ◽  
Simultaneous Formation ◽  
Lipid Ii ◽  
Target Binding ◽  
Dynamics Simulations ◽  
Similar Conformation ◽  
Insight Into

Lanthionine-containing peptides (lantibiotics) have been considered as pharmaceutical candidates for decades, although their clinical application has been restricted. Most lantibiotics kill bacteria via targeting and segregating of the cell wall precursor—membrane-inserted lipid II molecule—in some cases accompanied by pores formation. Nisin-like lantibiotics specifically bind to pyrophosphate (PPi) moiety of lipid II with their structurally similar N-terminal thioether rings A and B. Although possessing higher pore-forming capability, nisin, in some cases, is 10-fold less efficient in vivo as compared to related epidermin and gallidermin peptides, differing just in a few amino acid residues within their target-binding regions. Here, using molecular dynamics simulations, we investigated atomistic details of intermolecular interactions between the truncated analogues of these peptides (residues 1–12) and lipid II mimic (dimethyl pyrophosphate, DMPPi). The peptides adopt similar conformation upon DMPPi binding with backbone amide protons orienting into a single center capturing PPi moiety via simultaneous formation of up to seven hydrogen bonds. Epidermin and gallidermin adopt the complex-forming conformation twice as frequent as nisin does, enhancing the binding by the lysine 4 side chain. Introduction of the similar residue to nisin in silico improves the binding, providing ideas for further design of prototypic antibiotics.

scholarly journals Enzymatic Switching Between Archaeal DNA Polymerases Facilitates Abasic Site Bypass

2021 ◽  
Vol 12 ◽  
Author(s):  
Xu Feng ◽  
Baochang Zhang ◽  
Ruyi Xu ◽  
Zhe Gao ◽  
Xiaotong Liu ◽  
...  
Keyword(s):  
Dna Synthesis ◽  
Enzyme Kinetics ◽  
Dna Polymerases ◽  
Dna Lesions ◽  
Abasic Site ◽  
Lesion Bypass ◽  
Translesion Dna Synthesis ◽  
Kinetics Studies ◽  
Abasic Sites ◽  
Y Family

Abasic sites are among the most abundant DNA lesions encountered by cells. Their replication requires actions of specialized DNA polymerases. Herein, two archaeal specialized DNA polymerases were examined for their capability to perform translesion DNA synthesis (TLS) on the lesion, including Sulfolobuss islandicus Dpo2 of B-family, and Dpo4 of Y-family. We found neither Dpo2 nor Dpo4 is efficient to complete abasic sites bypass alone, but their sequential actions promote lesion bypass. Enzyme kinetics studies further revealed that the Dpo4’s activity is significantly inhibited at +1 to +3 site past the lesion, at which Dpo2 efficiently extends the primer termini. Furthermore, their activities are inhibited upon synthesis of 5–6 nt TLS patches. Once handed over to Dpo1, these substrates basically inactivate its exonuclease, enabling the transition from proofreading to polymerization of the replicase. Collectively, by functioning as an “extender” to catalyze further DNA synthesis past the lesion, Dpo2 bridges the activity gap between Dpo4 and Dpo1 in the archaeal TLS process, thus achieving more efficient lesion bypass.

Human DNA Polymerases λ and β Show Different Efficiencies of Translesion DNA Synthesis past Abasic Sites and Alternative Mechanisms for Frameshift Generation†

Biochemistry ◽  
2004 ◽  
Vol 43 (36) ◽  
pp. 11605-11615 ◽  
Author(s):  
Giuseppina Blanca ◽  
Giuseppe Villani ◽  
Igor Shevelev ◽  
Kristijan Ramadan ◽  
Silvio Spadari ◽  
...  
Keyword(s):  
Dna Synthesis ◽  
Dna Polymerases ◽  
Translesion Dna Synthesis ◽  
Abasic Sites ◽  
Human Dna ◽  
Translesion Dna
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