Characterization of the Entire Cystatin Gene Family in Barley and Their Target Cathepsin L-Like Cysteine-Proteases, Partners in the Hordein Mobilization during Seed Germination

Cathepsin L genes, which belonged to cysteine proteases, were a series of multifunctional protease and played important roles in a lot of pathological and physiological processes. In this study, we analyzed the characteristics a cathepsin L (named Mn-CL2) in the female oriental river prawn, Macrobrachium nipponense which was involved in ovary maturation. The Mn-CL2 was1,582 bp in length, including a 978 bp open reading frame that encoded 326 amino acids. The Mn-CL2 was classified into the cathepsin L group by phylogenetic analysis. Real-time PCR (qPCR) analysis indicated that Mn-CL2 was highly expressed in the hepatopancreas and ovaries of female prawns. During the different ovarian stages, Mn-CL2 expression in the hepatopancreas and ovaries peaked before ovarian maturation. In situ hybridization studies revealed that Mn-CL2 was localized in the oocyte of the ovary. Injection of Mn-CL2 dsRNA significantly reduced the expression of vitellogenin. Changes in the gonad somatic index also confirmed the inhibitory effects of Mn-CL2 dsRNA on ovary maturation. These results suggest that Mn-CL2 has a key role in promoting ovary maturation.

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Bioinformatics analysis and characterization of a secretory cystatin from Thelohanellus kitauei

10.21203/rs.3.rs-26747/v2 ◽

2020 ◽

Author(s):

Fengli Zhang ◽

Yalin Yang ◽

Chenchen Gao ◽

Yuanyuan Yao ◽

Rui Xia ◽

...

Keyword(s):

Binding Energy ◽

Inhibitory Activity ◽

Cathepsin B ◽

Cathepsin L ◽

Cysteine Proteases ◽

Economic Losses ◽

Amino Acid Residues ◽

Gene Encoding ◽

Multiple Sequence ◽

Cystatin Gene

Abstract Thelohanellus kitauei , is a group of obligate parasitic Myxozoans, which causes intestinal giant-cystic disease of common carp ( Cyprinus carpio) and has resulted in significant economic losses in carp farms. Cystatin secreted by parasites can regulate the immune response of host to facilitate parasite’s survival. In this study, the secretory TK-cystatin gene, encoding a protein of 120 amino acid residues (13.65 kDa), was cloned from T. kitauei genome. Phylogenetic analysis showed that TK-cystatin gene is closely related to the cystatin-A from Hydra vulgaris . Multiple sequence alignment revealed that TK-cystatin had three conserved motifs: N-terminal G 19 G 20 , Q 73 VVAG 77 , and C-terminal L 102 P 103 . Molecular docking between TK-cystatin and three cysteine proteases showed a lower binding energy (-13 kal/mol) with cathepsin L whereas a higher binding energy (-8.6 kal/mol) with cathepsin B. TK-cystatin gene was expressed in Escherichia coli . Activity assays revealed that TK-cystatin has stronger inhibitory activity on endopeptidases (papain and cathepsin L) and weaker inhibitory activity on exopeptidase (cathepsin B). TK-cystatin was stable under the condition of acidity or alkalinity or below 57 °C. This study laid a foundation for the design and development of the anti-TK-cystatin vaccine in carp culture in the future.

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Bioinformatics analysis and characterization of a secretory cystatin from Thelohanellus kitauei

10.21203/rs.3.rs-26747/v1 ◽

2020 ◽

Author(s):

Fengli Zhang ◽

Yalin Yang ◽

Chenchen Gao ◽

Yuanyuan Yao ◽

Rui Xia ◽

...

Keyword(s):

Binding Energy ◽

Inhibitory Activity ◽

Cathepsin B ◽

Cathepsin L ◽

Cysteine Proteases ◽

Economic Losses ◽

Amino Acid Residues ◽

Gene Encoding ◽

Multiple Sequence ◽

Cystatin Gene

Abstract Thelohanellus kitauei, is a group of obligate parasitic Myxozoans, which causes intestinal giant-cystic disease of common carp (Cyprinus carpio) and has resulted in significant economic losses in carp farms. Cystatin secreted by parasites can regulate the immune response of host to facilitate parasite’s survival. In this study, the secretory TK-cystatin gene, encoding a protein of 120 amino acid residues (13.65 kDa), was cloned from T. kitauei genome. Phylogenetic analysis showed that TK-cystatin gene is closely related to the cystatin-A from Hydra vulgaris. Multiple sequence alignment revealed that TK-cystatin had three conserved motifs: N-terminal G19G20, Q73VVAG77, and C-terminal L102P103. Molecular docking between TK-cystatin and three cysteine proteases showed a lower binding energy (-13 kal/mol) with cathepsin L whereas a higher binding energy (-8.6 kal/mol) with cathepsin B. TK-cystatin gene was expressed in Escherichia coli. Activity assays revealed that TK-cystatin has stronger inhibitory activity on endopeptidases (papain and cathepsin L) and weaker inhibitory activity on exopeptidase (cathepsin B). TK-cystatin was stable under the condition of acidity or alkalinity or below 57 °C. This study laid a foundation for the design and development of the anti-TK-cystatin vaccine in carp culture in the future.

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Production and Characterization of Two Variants of Human Cystatin SA Encoded by Two Alleles at theCST2Locus of the Type 2 Cystatin Gene Family

Archives of Biochemistry and Biophysics ◽

10.1006/abbi.1997.0609 ◽

1998 ◽

Vol 352 (2) ◽

pp. 199-206 ◽

Cited By ~ 17

Author(s):

Eiichi Saitoh ◽

Kiyoshi Minaguchi ◽

Osamu Ishibashi

Keyword(s):

Gene Family ◽

Cystatin Gene ◽

Human Cystatin

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Expression of beta-galactosidase multiple forms during barley (Hordeum vulgare) seed germination. Separation and characterization of enzyme isoforms

Physiologia Plantarum ◽

10.1034/j.1399-3054.1991.820313.x ◽

1991 ◽

Vol 82 (3) ◽

pp. 413-418 ◽

Cited By ~ 3

Author(s):

Thomas Giannakouros ◽

Athanasios Karagiorgos ◽

George Simos

Keyword(s):

Hordeum Vulgare ◽

Seed Germination ◽

Multiple Forms ◽

Enzyme Isoforms ◽

Beta Galactosidase

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Whole-Genome Sequence Characterization of Primary Auxin-Responsive Aux/IAA Gene Family in Sorghum (Sorghum bicolor L.)

ACTA AGRONOMICA SINICA ◽

10.3724/sp.j.1006.2010.00688 ◽

2010 ◽

Vol 36 (4) ◽

pp. 688-694

Author(s):

Yi-Jun WANG ◽

Yan-Ping LÜ ◽

Qin XIE ◽

De-Xiang DENG ◽

Yun-Long BIAN

Keyword(s):

Sorghum Bicolor ◽

Gene Family ◽

Genome Sequence ◽

Whole Genome Sequence ◽

Whole Genome ◽

Sequence Characterization ◽

I Gene

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In Silico Identification and Molecular Characterization of Extracellular Cathepsin L Proteases from Giardia duodenalis

Current Proteomics ◽

10.2174/1570164617666191016170628 ◽

2020 ◽

Vol 17 (4) ◽

pp. 342-351

Author(s):

Sergio A. Durán-Pérez ◽

José G. Rendón-Maldonado ◽

Lucio de Jesús Hernandez-Diaz ◽

Annete I. Apodaca-Medina ◽

Maribel Jiménez-Edeza ◽

...

Keyword(s):

In Silico ◽

Cathepsin L ◽

Three Dimensional ◽

Giardia Duodenalis ◽

Evolutionary Genetics ◽

Extracellular Proteins ◽

In Silico Identification ◽

Dimensional Models ◽

Three Dimensional Models

Background: The protozoan Giardia duodenalis, which causes giardiasis, is an intestinal parasite that commonly affects humans, mainly pre-school children. Although there are asymptomatic cases, the main clinical features are chronic and acute diarrhea, nausea, abdominal pain, and malabsorption syndrome. Little is currently known about the virulence of the parasite, but some cases of chronic gastrointestinal alterations post-infection have been reported even when the infection was asymptomatic, suggesting that the cathepsin L proteases of the parasite may be involved in the damage at the level of the gastrointestinal mucosa. Objective: The aim of this study was the in silico identification and characterization of extracellular cathepsin L proteases in the proteome of G. duodenalis. Methods: The NP_001903 sequence of cathepsin L protease from Homo sapienswas searched against the Giardia duodenalisproteome. The subcellular localization of Giardia duodenaliscathepsin L proteases was performed in the DeepLoc-1.0 server. The construction of a phylogenetic tree of the extracellular proteins was carried out using the Molecular Evolutionary Genetics Analysis software (MEGA X). The Robetta server was used for the construction of the three-dimensional models. The search for possible inhibitors of the extracellular cathepsin L proteases of Giardia duodenaliswas performed by entering the three-dimensional structures in the FINDSITEcomb drug discovery tool. Results: Based on the amino acid sequence of cathepsin L from Homo sapiens, 8 protein sequences were identified that have in their modular structure the Pept_C1A domain characteristic of cathepsins and two of these proteins (XP_001704423 and XP_001704424) are located extracellularly. Threedimensional models were designed for both extracellular proteins and several inhibitory ligands with a score greater than 0.9 were identified. In vitrostudies are required to corroborate if these two extracellular proteins play a role in the virulence of Giardia duodenalisand to discover ligands that may be useful as therapeutic targets that interfere in the mechanism of pathogenesis generated by the parasite. Conclusion: In silicoanalysis identified two proteins in the Giardia duodenalisprotein repertoire whose characteristics allowed them to be classified as cathepsin L proteases, which may be secreted into the extracellular medium to act as virulence factors. Three-dimensional models of both proteins allowed the identification of inhibitory ligands with a high score. The results suggest that administration of those compounds might be used to block the endopeptidase activity of the extracellular cathepsin L proteases, interfering with the mechanisms of pathogenesis of the protozoan parasite Giardia duodenalis.

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