scholarly journals Rationally Modified Antimicrobial Peptides from the N-Terminal Domain of Human RNase 3 Show Exceptional Serum Stability

2021 ◽  
Author(s):  
Daniel Sandín ◽  
Javier Valle ◽  
Belén Chaves-Arquero ◽  
Guillem Prats-Ejarque ◽  
María Nieves Larrosa ◽  
...  
Keyword(s):  
Antimicrobial Peptides ◽  
Serum Stability ◽  
Terminal Domain ◽  
Human Rnase

scholarly journals Easy Strategy To Protect Antimicrobial Peptides from Fast Degradation in Serum

2010 ◽  
Vol 54 (9) ◽  
pp. 4003-4005 ◽  
Author(s):  
Daniel Knappe ◽  
Petra Henklein ◽  
Ralf Hoffmann ◽  
Kai Hilpert
Keyword(s):  
Antimicrobial Activity ◽  
Drug Development ◽  
Antimicrobial Peptides ◽  
Propionic Acid ◽  
Mouse Serum ◽  
Development Programs ◽  
Serum Stability ◽  
Arginine Residues ◽  
Low Serum

ABSTRACT Antimicrobial peptides are promising novel peptide leads, but their low serum stability often limits their further consideration in drug development programs. Here, we describe a generally applicable strategy to stabilize peptides against serum proteases by replacing arginine residues with α-amino-3-guanidino-propionic acid (Agp). Peptide NH2-RRWRIVVIRVRR-CONH2 was nearby totally degraded after 8 h in mouse serum, whereas the variant with Agp substituted was degraded less than 20%. The antimicrobial activity was not affected.

scholarly journals Expression of human ACE2 N-terminal domain, part of the receptor for SARS-CoV-2, in fusion with maltose binding protein, E. coli ribonuclease I and human RNase A

2021 ◽  
Author(s):  
Shuang-yong Xu ◽  
Alexey Fomenkov ◽  
Tien-Hao Chen ◽  
Erbay Yigit
Keyword(s):  
Viral Genome ◽  
Antiviral Agents ◽  
Cell Receptor ◽  
Rna Degradation ◽  
Rnase A ◽  
Rnase Iii ◽  
E Coli ◽  
Terminal Domain ◽  
Human Rnase

AbstractThe SARS-CoV-2 viral genome contains a positive-strand single-stranded RNA of ~30 kb. Human ACE2 protein is the receptor for SARS-CoV-2 virus attachment and initiation of infection. We propose to use ribonucleases (RNases) as antiviral agents to destroy the viral genome in vitro. In the virions the RNA is protected by viral capsid proteins, membrane proteins and nucleocapsid proteins. To overcome this protection we set out to construct RNase fusion with human ACE2 receptor N-terminal domain (ACE2NTD). We constructed six proteins expressed in E. coli cells: 1) MBP-ACE2NTD, 2) ACE2NTD-GFP, 3) RNase I (6xHis), 4) RNase III (6xHis), 5) RNase I-ACE2NTD (6xHis), and 6) human RNase A-ACE2NTD150 (6xHis). We evaluated fusion expression in different E. coli strains, partially purified MBP-ACE2NTD protein from the soluble fraction of bacterial cell lysate, and refolded MBP-ACE2NTD protein from inclusion body. The engineered RNase I-ACE2NTD (6xHis) and hRNase A-ACE2NTD (6xHis) fusions are active in cleaving COVID-19 RNA in vitro. The recombinant RNase I (6xHis) and RNase III (6xHis) are active in cleaving RNA and dsRNA in test tube. This study provides a proof-of-concept for construction of fusion protein between human cell receptor and nuclease that may be used to degrade viral nucleic acids in our environment.Graphical AbstractCartoon illustration part of this work (Human ACE2 N-terminal domain tethered to RNase A and RNA degradation by the fusion enzyme).

scholarly journals Ultrashort Antimicrobial Peptides with Antiendotoxin Properties

2015 ◽  
Vol 59 (8) ◽  
pp. 5052-5056 ◽  
Author(s):  
Ya-Han Chih ◽  
Yen-Shan Lin ◽  
Bak-Sau Yip ◽  
Hsiu-Ju Wei ◽  
Hung-Lun Chu ◽  
...  
Keyword(s):  
Immune System ◽  
Antimicrobial Peptides ◽  
Salt Resistance ◽  
Host Immune System ◽  
Serum Stability ◽  
Proinflammatory Responses ◽  
Stimulation Of

ABSTRACTRelease of lipopolysaccharide (LPS) (endotoxin) from bacteria into the bloodstream may cause serious unwanted stimulation of the host immune system. Some but not all antimicrobial peptides can neutralize LPS-stimulated proinflammatory responses. Salt resistance and serum stability of short antimicrobial peptides can be boosted by adding β-naphthylalanine to their termini. Herein, significant antiendotoxin effects were observedin vitroandin vivowith the β-naphthylalanine end-tagged variants of the short antimicrobial peptides S1 and KWWK.

scholarly journals Boosting Salt Resistance of Short Antimicrobial Peptides

2013 ◽  
Vol 57 (8) ◽  
pp. 4050-4052 ◽  
Author(s):  
Hung-Lun Chu ◽  
Hui-Yuan Yu ◽  
Bak-Sau Yip ◽  
Ya-Han Chih ◽  
Chong-Wen Liang ◽  
...  
Keyword(s):  
Amino Acid ◽  
Antimicrobial Peptides ◽  
Salt Resistance ◽  
High Salt ◽  
Serum Stability ◽  
Pharmaceutical Agents

ABSTRACTThe efficacies of many antimicrobial peptides are greatly reduced under high salt concentrations, therefore limiting their use as pharmaceutical agents. Here, we describe a strategy to boost salt resistance and serum stability of short antimicrobial peptides by adding the nonnatural bulky amino acid β-naphthylalanine to their termini. The activities of the short salt-sensitive tryptophan-rich peptide S1 were diminished at high salt concentrations, whereas the activities of its β-naphthylalanine end-tagged variants were less affected.

scholarly journals SppI Forms a Membrane Protein Complex with SppA and Inhibits Its Protease Activity in Bacillus subtilis

mSphere ◽  
2020 ◽  
Vol 5 (5) ◽  
Author(s):  
Gabriela Henriques ◽  
Stephen McGovern ◽  
Jolanda Neef ◽  
Minia Antelo-Varela ◽  
Friedrich Götz ◽  
...  
Keyword(s):  
Bacillus Subtilis ◽  
Membrane Protein ◽  
Antimicrobial Peptides ◽  
Protease Activity ◽  
Cationic Antimicrobial Peptide ◽  
Content Type ◽  
Terminal Domain ◽  
Increased Sensitivity

ABSTRACT The membrane protease SppA of Bacillus subtilis was first described as a signal peptide peptidase and later shown to confer resistance to lantibiotics. Here, we report that SppA forms octameric complexes with YteJ, a membrane protein of thus-far-unknown function. Interestingly, sppA and yteJ deletion mutants exhibited no protein secretion defects. However, these mutant strains differed significantly in their resistance to antimicrobial peptides. In particular, sppA mutant cells displayed increased sensitivity to the lantibiotics nisin and subtilin and the human lysozyme-derived cationic antimicrobial peptide LP9. Importantly, YteJ was shown to antagonize SppA activity both in vivo and in vitro, and this SppA-inhibitory activity involved the C-terminal domain of YteJ, which was therefore renamed SppI. Most likely, SppI-mediated control is needed to protect B. subtilis against the potentially detrimental protease activity of SppA since a mutant overexpressing sppA by itself displayed defects in cell division. Altogether, we conclude that the SppA-SppI complex of B. subtilis has a major role in protection against antimicrobial peptides. IMPORTANCE Our study presents new insights into the molecular mechanism that regulates the activity of SppA, a widely conserved bacterial membrane protease. We show that the membrane proteins SppA and SppI form a complex in the Gram-positive model bacterium B. subtilis and that SppI inhibits SppA protease activity in vitro and in vivo. Furthermore, we demonstrate that the C-terminal domain of SppI is involved in SppA inhibition. Since SppA, through its protease activity, contributes directly to resistance to lantibiotic peptides and cationic antibacterial peptides, we propose that the conserved SppA-SppI complex could play a major role in the evasion of bactericidal peptides, including those produced as part of human innate immune defenses.

Structure of clathrin cages and coats in ice

1986 ◽  
Vol 44 ◽  
pp. 94-97
Author(s):  
G.P.A. Vigers ◽  
R.A. Crowther ◽  
B.M.F. Pearse
Keyword(s):  
Electron Microscopy ◽  
Heavy Chain ◽  
Outer Part ◽  
Coated Vesicles ◽  
Eukaryotic Cells ◽  
Coat Proteins ◽  
Terminal Domain ◽  
Clathrin Heavy Chain ◽  
Membrane Components

Clathrin forms the polyhedral cage of coated vesicles, which mediate the transfer of selected membrane components within eukaryotic cells. Clathrin cages and coated vesicles have been extensively studied by electron microscopy of negatively stained preparations and shadowed specimens. From these studies the gross morphology of the outer part of the polyhedral coat has been established and some features of the packing of clathrin trimers into the coat have also been described. However these previous studies have not revealed any internal details about the position of the terminal domain of the clathrin heavy chain, the location of the 100kd-50kd accessory coat proteins or the interactions of the coat with the enclosed membrane.

Crystal structures of Magnaporthe oryzae trehalose-6-phosphate synthase (MoTps1) suggest a model for catalytic process of Tps1

2019 ◽  
Vol 476 (21) ◽  
pp. 3227-3240 ◽  
Author(s):  
Shanshan Wang ◽  
Yanxiang Zhao ◽  
Long Yi ◽  
Minghe Shen ◽  
Chao Wang ◽  
...  
Keyword(s):  
Crystal Structures ◽  
Conformational Changes ◽  
Magnaporthe Oryzae ◽  
Structural Information ◽  
Complex Structure ◽  
Critical Role ◽  
Catalytic Process ◽  
Structural Basis ◽  
Salt Bridges ◽  
Terminal Domain

Trehalose-6-phosphate (T6P) synthase (Tps1) catalyzes the formation of T6P from UDP-glucose (UDPG) (or GDPG, etc.) and glucose-6-phosphate (G6P), and structural basis of this process has not been well studied. MoTps1 (Magnaporthe oryzae Tps1) plays a critical role in carbon and nitrogen metabolism, but its structural information is unknown. Here we present the crystal structures of MoTps1 apo, binary (with UDPG) and ternary (with UDPG/G6P or UDP/T6P) complexes. MoTps1 consists of two modified Rossmann-fold domains and a catalytic center in-between. Unlike Escherichia coli OtsA (EcOtsA, the Tps1 of E. coli), MoTps1 exists as a mixture of monomer, dimer, and oligomer in solution. Inter-chain salt bridges, which are not fully conserved in EcOtsA, play primary roles in MoTps1 oligomerization. Binding of UDPG by MoTps1 C-terminal domain modifies the substrate pocket of MoTps1. In the MoTps1 ternary complex structure, UDP and T6P, the products of UDPG and G6P, are detected, and substantial conformational rearrangements of N-terminal domain, including structural reshuffling (β3–β4 loop to α0 helix) and movement of a ‘shift region' towards the catalytic centre, are observed. These conformational changes render MoTps1 to a ‘closed' state compared with its ‘open' state in apo or UDPG complex structures. By solving the EcOtsA apo structure, we confirmed that similar ligand binding induced conformational changes also exist in EcOtsA, although no structural reshuffling involved. Based on our research and previous studies, we present a model for the catalytic process of Tps1. Our research provides novel information on MoTps1, Tps1 family, and structure-based antifungal drug design.

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