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16 pages, 2773 KiB

Open AccessArticle

Svx Peptidases of Phytopathogenic Pectolytic Bacteria: Structural, Catalytic and Phytoimmune Properties

by Natalia Tendiuk, Anastasiya Diakonova, Olga Petrova, Timur Mukhametzyanov, Olga Makshakova and Vladimir Gorshkov

Int. J. Mol. Sci. 2024, 25(2), 756; https://doi.org/10.3390/ijms25020756 - 7 Jan 2024

Viewed by 1176

Abstract

Svx proteins are virulence factors secreted by phytopathogenic bacteria of the Pectobacterium genus into the host plant cell wall. Svx-encoding genes are present in almost all species of the soft rot Pectobacteriaceae (Pectobacterium and Dickeya genera). The Svx of P. atrosepticum ( [...] Read more.

Svx proteins are virulence factors secreted by phytopathogenic bacteria of the Pectobacterium genus into the host plant cell wall. Svx-encoding genes are present in almost all species of the soft rot Pectobacteriaceae (Pectobacterium and Dickeya genera). The Svx of P. atrosepticum (Pba) has been shown to be a gluzincin metallopeptidase that presumably targets plant extensins, proteins that contribute to plant cell wall rigidity and participate in cell signaling. However, the particular “output” of the Pba Svx action in terms of plant-pathogen interactions and plant immune responses remained unknown. The Svx proteins are largely unexplored in Dickeya species, even though some of them have genes encoding two Svx homologs. Therefore, our study aims to compare the structural and catalytic properties of the Svx proteins of Pba and D. solani (Dso) and to test the phytoimmune properties of these proteins. Two assayed Dso Svx proteins, similar to Pba Svx, were gluzincin metallopeptidases with conservative tertiary structures. The two domains of the Svx proteins form electronegative clefts where the active centers of the peptidase domains are located. All three assayed Svx proteins possessed phytoimmunosuppressory properties and induced ethylene-mediated plant susceptible responses that play a decisive role in Pba-caused disease. Full article

(This article belongs to the Special Issue Microbial Proteases: Structure, Function and Role in Pathogenesis)

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11 pages, 2439 KiB

Open AccessArticle

Hemostatically Active Proteinase Produced by Aspergillus ochraceus: Key Specific Properties and Effect on Target Proteins

by Alexander A. Osmolovskiy and Valeriana G. Kreyer

Int. J. Mol. Sci. 2023, 24(18), 13870; https://doi.org/10.3390/ijms241813870 - 8 Sep 2023

Cited by 1 | Viewed by 782

Abstract

The effect of A. ochraceus proteinase on the proteins of the human hemostasis system, fibrin, fibrinogen, plasminogen, protein C, and factor X, was studied. These proteins are key targets for proteolytic enzymes in therapy and diagnosis of thromboembolic complications. It was shown that [...] Read more.

The effect of A. ochraceus proteinase on the proteins of the human hemostasis system, fibrin, fibrinogen, plasminogen, protein C, and factor X, was studied. These proteins are key targets for proteolytic enzymes in therapy and diagnosis of thromboembolic complications. It was shown that A. ochraceus proteinase efficiently cleaves fibrin and fibrinogen, but does not act precisely, since it cuts all three subunits of these proteins. The proteinase did not have an activating effect on the plasminogen, a precursor of plasminogen and plasmin. The proteinase of A. ochraceus was shown to be the first fungal proteinase with proven activating activity towards the human hemostasis system factors protein C and factor X. For protein C activation, A. ochraceus proteinase requires Ca²⁺ ions. The enzyme was found to be sensitive to thrombin inhibitors, but not to plasmin inhibitors. A proteolytic action profile of the scope of this proteinase as a proteinase with activating protein C, factor X, and plasmin-like activity was proposed. Full article

(This article belongs to the Special Issue Microbial Proteases: Structure, Function and Role in Pathogenesis)

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19 pages, 1505 KiB

Open AccessArticle

Transcriptomic Analysis Followed by the Isolation of Extracellular Bacteriolytic Proteases from Lysobacter capsici VKM B-2533^T

by Alexey Afoshin, Irina Kudryakova, Sergey Tarlachkov, Elena Leontyevskaya, Dmitry Zelenov, Pavel Rudenko and Natalya Leontyevskaya (Vasilyeva)

Int. J. Mol. Sci. 2023, 24(14), 11652; https://doi.org/10.3390/ijms241411652 - 19 Jul 2023

Cited by 4 | Viewed by 1195

Abstract

The aim of the study was to search for, isolate and characterize new bacteriolytic enzymes that show promising potential for their use in medicine, agriculture and veterinary. Using a transcriptomic analysis, we annotated in Lysobacter capsici VKM B-2533^T the genes of known [...] Read more.

The aim of the study was to search for, isolate and characterize new bacteriolytic enzymes that show promising potential for their use in medicine, agriculture and veterinary. Using a transcriptomic analysis, we annotated in Lysobacter capsici VKM B-2533^T the genes of known bacteriolytic and antifungal enzymes, as well as of antibiotics, whose expression levels increased when cultivated on media conducive to the production of antimicrobial agents. The genes of the secreted putative bacteriolytic proteases were also annotated. Two new bacteriolytic proteases, Serp and Serp3, were isolated and characterized. The maximum bacteriolytic activities of Serp and Serp3 were exhibited at low ionic strength of 10 mM Tris-HCl, and high temperatures of, respectively, 80 °C and 70 °C. The pH optimum for Serp was 8.0; for Serp3, it was slightly acidic, at 6.0. Both enzymes hydrolyzed autoclaved cells of Micrococcus luteus Ac-2230^T, Proteus vulgaris H-19, Pseudomonas aeruginosa and Staphylococcus aureus 209P. Serp also digested cells of Bacillus cereus 217. Both enzymes hydrolyzed casein and azofibrin. The newly discovered enzymes are promising for developing proteolytic antimicrobial drugs on their basis. Full article

(This article belongs to the Special Issue Microbial Proteases: Structure, Function and Role in Pathogenesis)

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18 pages, 4335 KiB

Open AccessArticle

Characteristics and Application of a Novel Cold-Adapted and Salt-Tolerant Protease EK4-1 Produced by an Arctic Bacterium Mesonia algae K4-1

by Hailian Rao, Ran Huan, Yidan Chen, Xun Xiao, Wenzhao Li and Hailun He

Int. J. Mol. Sci. 2023, 24(9), 7985; https://doi.org/10.3390/ijms24097985 - 28 Apr 2023

Cited by 3 | Viewed by 1416

Abstract

Mesonia algae K4-1 from the Arctic secretes a novel cold-adapted and salt-tolerant protease EK4-1. It has the highest sequence similarity with Stearolysin, an M4 family protease from Geobacillus stearothermophilus, with only 45% sequence identity, and is a novel M4 family protease. Ek4-1 [...] Read more.

Mesonia algae K4-1 from the Arctic secretes a novel cold-adapted and salt-tolerant protease EK4-1. It has the highest sequence similarity with Stearolysin, an M4 family protease from Geobacillus stearothermophilus, with only 45% sequence identity, and is a novel M4 family protease. Ek4-1 has a low optimal catalytic temperature (40 °C) and is stable at low temperatures. Moreover, EK4-1 is still active in 4 mol/L NaCl solution and is tolerant to surfactants, oxidizing agents and organic solvents; furthermore, it prefers the hydrolysis of peptide bonds at the P1’ position as the hydrophobic residues, such as Leu, Phe and Val, and amino acids with a long side chain, such as Phe and Tyr. Mn²⁺and Mg²⁺ significantly promoted enzyme activity, while Fe³⁺, Co⁺, Zn²⁺ and Cu²⁺ significantly inhibited enzyme activity. Amino acid composition analysis showed that EK4-1 had more small-side-chain amino acids and fewer large-side-chain amino acids. Compared with a thermophilic protease Stearolysin, the cold-adapted protease EK4-1 contains more random coils (48.07%) and a larger active pocket (727.42 Å³). In addition, the acidic amino acid content of protease EK4-1 was higher than that of the basic amino acid, which might be related to the salt tolerance of protease. Compared with the homologous proteases EB62 and E423, the cold-adapted protease EK4-1 was more efficient in the proteolysis of grass carp skin, salmon skin and casein at a low temperature, and produced a large number of antioxidant peptides, with DPPH, ·OH and ROO· scavenging activities. Therefore, cold-adapted and salt-tolerant protease EK4-1 offers wide application prospects in the cosmetic and detergent industries. Full article

(This article belongs to the Special Issue Microbial Proteases: Structure, Function and Role in Pathogenesis)

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Review

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12 pages, 1597 KiB

Open AccessReview

The Secreted Aminopeptidase of Pseudomonas aeruginosa (PaAP)

by Efrat Kessler

Int. J. Mol. Sci. 2024, 25(15), 8444; https://doi.org/10.3390/ijms25158444 - 2 Aug 2024

Viewed by 275

Abstract

Pseudomonas aeruginosa is an opportunistic pathogen that causes severe infections in compromised hosts. P. aeruginosa infections are difficult to treat because of the inherent ability of the bacteria to develop antibiotic resistance, secrete a variety of virulence factors, and form biofilms. The secreted [...] Read more.

Pseudomonas aeruginosa is an opportunistic pathogen that causes severe infections in compromised hosts. P. aeruginosa infections are difficult to treat because of the inherent ability of the bacteria to develop antibiotic resistance, secrete a variety of virulence factors, and form biofilms. The secreted aminopeptidase (PaAP) is an emerging virulence factor, key in providing essential low molecular weight nutrients and a cardinal modulator of biofilm development. PaAP is therefore a new potential target for therapy of P. aeruginosa infections. The present review summarizes the current knowledge of PaAP, with special emphasis on its biochemical and enzymatic properties, activation mechanism, biological roles, regulation, and structure. Recently developed specific inhibitors and their potential as adjuncts in the treatment of P. aeruginosa infections are also described. Full article

(This article belongs to the Special Issue Microbial Proteases: Structure, Function and Role in Pathogenesis)

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14 pages, 7704 KiB

Open AccessReview

The Balance between Protealysin and Its Substrate, the Outer Membrane Protein OmpX, Regulates Serratia proteamaculans Invasion

by Olga Tsaplina

Int. J. Mol. Sci. 2024, 25(11), 6159; https://doi.org/10.3390/ijms25116159 - 3 Jun 2024

Viewed by 372

Abstract

Serratia are opportunistic bacteria, causing infections in plants, insects, animals and humans under certain conditions. The development of bacterial infection in the human body involves several stages of host–pathogen interaction, including entry into non-phagocytic cells to evade host immune cells. The facultative pathogen [...] Read more.

Serratia are opportunistic bacteria, causing infections in plants, insects, animals and humans under certain conditions. The development of bacterial infection in the human body involves several stages of host–pathogen interaction, including entry into non-phagocytic cells to evade host immune cells. The facultative pathogen Serratia proteamaculans is capable of penetrating eukaryotic cells. These bacteria synthesize an actin-specific metalloprotease named protealysin. After transformation with a plasmid carrying the protealysin gene, noninvasive E. coli penetrate eukaryotic cells. This suggests that protealysin may play a key role in S. proteamaculans invasion. This review addresses the mechanisms underlying protealysin’s involvement in bacterial invasion, highlighting the main findings as follows. Protealysin can be delivered into the eukaryotic cell by the type VI secretion system and/or by bacterial outer membrane vesicles. By cleaving actin in the host cell, protealysin can mediate the reversible actin rearrangements required for bacterial invasion. However, inactivation of the protealysin gene leads to an increase, rather than decrease, in the intensity of S. proteamaculans invasion. This indicates the presence of virulence factors among bacterial protealysin substrates. Indeed, protealysin cleaves the virulence factors, including the bacterial surface protein OmpX. OmpX increases the expression of the EGFR and β1 integrin, which are involved in S. proteamaculans invasion. It has been shown that an increase in the invasion of genetically modified S. proteamaculans may be the result of the accumulation of full-length OmpX on the bacterial surface, which is not cleaved by protealysin. Thus, the intensity of the S. proteamaculans invasion is determined by the balance between the active protealysin and its substrate OmpX. Full article

(This article belongs to the Special Issue Microbial Proteases: Structure, Function and Role in Pathogenesis)

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25 pages, 3325 KiB

Open AccessReview

Secreted Aspartic Proteinases: Key Factors in Candida Infections and Host-Pathogen Interactions

by Grazyna Bras, Dorota Satala, Magdalena Juszczak, Kamila Kulig, Ewelina Wronowska, Aneta Bednarek, Marcin Zawrotniak, Maria Rapala-Kozik and Justyna Karkowska-Kuleta

Int. J. Mol. Sci. 2024, 25(9), 4775; https://doi.org/10.3390/ijms25094775 - 27 Apr 2024

Cited by 2 | Viewed by 1391

Abstract

Extracellular proteases are key factors contributing to the virulence of pathogenic fungi from the genus Candida. Their proteolytic activities are crucial for extracting nutrients from the external environment, degrading host defenses, and destabilizing the internal balance of the human organism. Currently, the [...] Read more.

Extracellular proteases are key factors contributing to the virulence of pathogenic fungi from the genus Candida. Their proteolytic activities are crucial for extracting nutrients from the external environment, degrading host defenses, and destabilizing the internal balance of the human organism. Currently, the enzymes most frequently described in this context are secreted aspartic proteases (Saps). This review comprehensively explores the multifaceted roles of Saps, highlighting their importance in biofilm formation, tissue invasion through the degradation of extracellular matrix proteins and components of the coagulation cascade, modulation of host immune responses via impairment of neutrophil and monocyte/macrophage functions, and their contribution to antifungal resistance. Additionally, the diagnostic challenges associated with Candida infections and the potential of Saps as biomarkers were discussed. Furthermore, we examined the prospects of developing vaccines based on Saps and the use of protease inhibitors as adjunctive therapies for candidiasis. Given the complex biology of Saps and their central role in Candida pathogenicity, a multidisciplinary approach may pave the way for innovative diagnostic strategies and open new opportunities for innovative clinical interventions against candidiasis. Full article

(This article belongs to the Special Issue Microbial Proteases: Structure, Function and Role in Pathogenesis)

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28 pages, 2830 KiB

Open AccessReview

Extracellular Cysteine Proteases of Key Intestinal Protozoan Pathogens—Factors Linked to Virulence and Pathogenicity

by Raúl Argüello-García, Julio César Carrero and M. Guadalupe Ortega-Pierres

Int. J. Mol. Sci. 2023, 24(16), 12850; https://doi.org/10.3390/ijms241612850 - 16 Aug 2023

Cited by 3 | Viewed by 1865

Abstract

Intestinal diseases caused by protistan parasites of the genera Giardia (giardiasis), Entamoeba (amoebiasis), Cryptosporidium (cryptosporidiosis) and Blastocystis (blastocystosis) represent a major burden in human and animal populations worldwide due to the severity of diarrhea and/or inflammation in susceptible hosts. These pathogens interact with [...] Read more.

Intestinal diseases caused by protistan parasites of the genera Giardia (giardiasis), Entamoeba (amoebiasis), Cryptosporidium (cryptosporidiosis) and Blastocystis (blastocystosis) represent a major burden in human and animal populations worldwide due to the severity of diarrhea and/or inflammation in susceptible hosts. These pathogens interact with epithelial cells, promoting increased paracellular permeability and enterocyte cell death (mainly apoptosis), which precede physiological and immunological disorders. Some cell-surface-anchored and molecules secreted from these parasites function as virulence markers, of which peptide hydrolases, particularly cysteine proteases (CPs), are abundant and have versatile lytic activities. Upon secretion, CPs can affect host tissues and immune responses beyond the site of parasite colonization, thereby increasing the pathogens’ virulence. The four intestinal protists considered here are known to secrete predominantly clan A (C1- and C2-type) CPs, some of which have been characterized. CPs of Giardia duodenalis (e.g., Giardipain-1) and Entamoeba histolytica (EhCPs 1-6 and EhCP112) degrade mucin and villin, cause damage to intercellular junction proteins, induce apoptosis in epithelial cells and degrade immunoglobulins, cytokines and defensins. In Cryptosporidium, five Cryptopains are encoded in its genome, but only Cryptopains 4 and 5 are likely secreted. In Blastocystis sp., a legumain-activated CP, called Blastopain-1, and legumain itself have been detected in the extracellular medium, and the former has similar adverse effects on epithelial integrity and enterocyte survival. Due to their different functions, these enzymes could represent novel drug targets. Indeed, some promising results with CP inhibitors, such as vinyl sulfones (K11777 and WRR605), the garlic derivative, allicin, and purified amoebic CPs have been obtained in experimental models, suggesting that these enzymes might be useful drug targets. Full article

(This article belongs to the Special Issue Microbial Proteases: Structure, Function and Role in Pathogenesis)

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