Expression of nitric oxide synthases from photosynthetic microorganisms improves growth and stress tolerance in E. coli

Autores
Correa Aragunde, Maria Natalia; Nejamkin, Andres; del Castello, Fiorella Paola; Foresi, Noelia Pamela; Lamattina, Lorenzo
Año de publicación
2020
Idioma
inglés
Tipo de recurso
documento de conferencia
Estado
versión publicada
Descripción
Nitric oxide synthase (NOS) catalyzes the oxidation of the substrate L-Arginine (Arg) to produce citrulline and nitric oxide (NO). We have characterized NOS from two photosynthetic microorganisms: the NOS from the alga Ostreococcus tauri (OtNOS) and the cyanobacteria Synechococcus PCC 7335 (SyNOS). OtNOS and SyNOS possess distinct biochemical properties. OtNOS is a canonical NOS similar to animal NOS with an ultrafast NO producing activity. On the contrary, an extra globin domain present in SyNOS enzyme oxidizes over 70 % of the NO-produced to nitrate (NO3-). Here we describethe expression of recombinant OtNOS and SyNOS in Escherichia coli BL12 strain and analyze bacterial growth and tolerance to nitrosative stress. Results show that the E. coli cultures expressing OtNOS and SyNOS reach a higher OD at the exponential phase with respect to bacteria transformed with the empty vector (EV). This result correlates with higher NOS protein levels assayed by immunoblot, total protein and nitrate content in NOS recombinant strain cultures. Moreover, the expression of SyNOS and at less extent of OtNOS confers the ability to grow in minimal medium with Arg as a sole N source (and plenty C-source), suggesting that NOS enzymes are active in E. coli. The high NO producing activity reported in OtNOS correlates with the flavohemoglobin hmp induction in E. coli strain expressing OtNOS, suggesting that this strain senses nitrosative stress. Furthermore, nitrosative stress generated by the addition of 1 mM of the NO donor sodium nitroprusside (SNP) reduced growth rate (0.4-fold respect to no SNP addition) in bacterial culture expressing EV. However, the expression of recombinant OtNOS and at less extent SyNOS, attenuated SNP toxicity (0.8- and 0.6-fold, respectively, compared to no SNP addition). E. coli does not synthesize the major NOS cofactor tetrahydrobiopterin (BH4). Bioinformatics tools and ligand docking analysis were used to provide evidence supporting tetrahydromonapterin (MH4) as a possible pterin cofactor required for NOS catalytic activity in E. coli. These results open an exciting new window about the versatility of pterin cofactor working in the different NOSs dispersed in distant organisms along the life tree. In summary, our results show that NOS from photosynthetic microorganisms increases the growth and confers nitrosative stress tolerance in E. coli. Supported by AGENCIA, CONICET and UNMdP
Fil: Correa Aragunde, Maria Natalia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina
Fil: Nejamkin, Andres. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina
Fil: del Castello, Fiorella Paola. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina
Fil: Foresi, Noelia Pamela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina
Fil: Lamattina, Lorenzo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina
Reunión Conjunta SAIB-SAMIGE 2020. LVI Reunión Anual de la Sociedad Argentina de Investigación en Bioquímica y Biología Molecular. LVI Reunión Anual Sociedad Argentina de Bioquímica y Biología Molecular (SAIB). XV Reunión Anual Sociedad Argentina de Microbiología General (SAMIGE)
Buenos Aires
Argentina
Sociedad Argentina de Investigación en Bioquímica y Biología Molecular
Asociación Civil de Microbiología General
Materia
BACTERIA
GROWTH
NITRIC OXIDE SYNTHASE
OXIDATIVE STRESS
Nivel de accesibilidad
acceso abierto
Condiciones de uso
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
Repositorio
CONICET Digital (CONICET)
Institución
Consejo Nacional de Investigaciones Científicas y Técnicas
OAI Identificador
oai:ri.conicet.gov.ar:11336/217194

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network_name_str CONICET Digital (CONICET)
spelling Expression of nitric oxide synthases from photosynthetic microorganisms improves growth and stress tolerance in E. coliCorrea Aragunde, Maria NataliaNejamkin, Andresdel Castello, Fiorella PaolaForesi, Noelia PamelaLamattina, LorenzoBACTERIAGROWTHNITRIC OXIDE SYNTHASEOXIDATIVE STRESShttps://purl.org/becyt/ford/1.6https://purl.org/becyt/ford/1Nitric oxide synthase (NOS) catalyzes the oxidation of the substrate L-Arginine (Arg) to produce citrulline and nitric oxide (NO). We have characterized NOS from two photosynthetic microorganisms: the NOS from the alga Ostreococcus tauri (OtNOS) and the cyanobacteria Synechococcus PCC 7335 (SyNOS). OtNOS and SyNOS possess distinct biochemical properties. OtNOS is a canonical NOS similar to animal NOS with an ultrafast NO producing activity. On the contrary, an extra globin domain present in SyNOS enzyme oxidizes over 70 % of the NO-produced to nitrate (NO3-). Here we describethe expression of recombinant OtNOS and SyNOS in Escherichia coli BL12 strain and analyze bacterial growth and tolerance to nitrosative stress. Results show that the E. coli cultures expressing OtNOS and SyNOS reach a higher OD at the exponential phase with respect to bacteria transformed with the empty vector (EV). This result correlates with higher NOS protein levels assayed by immunoblot, total protein and nitrate content in NOS recombinant strain cultures. Moreover, the expression of SyNOS and at less extent of OtNOS confers the ability to grow in minimal medium with Arg as a sole N source (and plenty C-source), suggesting that NOS enzymes are active in E. coli. The high NO producing activity reported in OtNOS correlates with the flavohemoglobin hmp induction in E. coli strain expressing OtNOS, suggesting that this strain senses nitrosative stress. Furthermore, nitrosative stress generated by the addition of 1 mM of the NO donor sodium nitroprusside (SNP) reduced growth rate (0.4-fold respect to no SNP addition) in bacterial culture expressing EV. However, the expression of recombinant OtNOS and at less extent SyNOS, attenuated SNP toxicity (0.8- and 0.6-fold, respectively, compared to no SNP addition). E. coli does not synthesize the major NOS cofactor tetrahydrobiopterin (BH4). Bioinformatics tools and ligand docking analysis were used to provide evidence supporting tetrahydromonapterin (MH4) as a possible pterin cofactor required for NOS catalytic activity in E. coli. These results open an exciting new window about the versatility of pterin cofactor working in the different NOSs dispersed in distant organisms along the life tree. In summary, our results show that NOS from photosynthetic microorganisms increases the growth and confers nitrosative stress tolerance in E. coli. Supported by AGENCIA, CONICET and UNMdPFil: Correa Aragunde, Maria Natalia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; ArgentinaFil: Nejamkin, Andres. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; ArgentinaFil: del Castello, Fiorella Paola. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; ArgentinaFil: Foresi, Noelia Pamela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; ArgentinaFil: Lamattina, Lorenzo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; ArgentinaReunión Conjunta SAIB-SAMIGE 2020. LVI Reunión Anual de la Sociedad Argentina de Investigación en Bioquímica y Biología Molecular. LVI Reunión Anual Sociedad Argentina de Bioquímica y Biología Molecular (SAIB). XV Reunión Anual Sociedad Argentina de Microbiología General (SAMIGE)Buenos AiresArgentinaSociedad Argentina de Investigación en Bioquímica y Biología MolecularAsociación Civil de Microbiología GeneralTech Science Press2020info:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/conferenceObjectCongresoJournalhttp://purl.org/coar/resource_type/c_5794info:ar-repo/semantics/documentoDeConferenciaapplication/pdfapplication/pdfapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/217194Expression of nitric oxide synthases from photosynthetic microorganisms improves growth and stress tolerance in E. coli; Reunión Conjunta SAIB-SAMIGE 2020. LVI Reunión Anual de la Sociedad Argentina de Investigación en Bioquímica y Biología Molecular. LVI Reunión Anual Sociedad Argentina de Bioquímica y Biología Molecular (SAIB). XV Reunión Anual Sociedad Argentina de Microbiología General (SAMIGE); Buenos Aires; Argentina; 2020; 108-1080327-95451667-5746CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/url/https://www.samige.org.ar/admin/news/files/170-TSP_BIOCELL_42376.pdfNacionalinfo:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-09-29T10:26:51Zoai:ri.conicet.gov.ar:11336/217194instacron:CONICETInstitucionalhttp://ri.conicet.gov.ar/Organismo científico-tecnológicoNo correspondehttp://ri.conicet.gov.ar/oai/requestdasensio@conicet.gov.ar; lcarlino@conicet.gov.arArgentinaNo correspondeNo correspondeNo correspondeopendoar:34982025-09-29 10:26:51.802CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv Expression of nitric oxide synthases from photosynthetic microorganisms improves growth and stress tolerance in E. coli
title Expression of nitric oxide synthases from photosynthetic microorganisms improves growth and stress tolerance in E. coli
spellingShingle Expression of nitric oxide synthases from photosynthetic microorganisms improves growth and stress tolerance in E. coli
Correa Aragunde, Maria Natalia
BACTERIA
GROWTH
NITRIC OXIDE SYNTHASE
OXIDATIVE STRESS
title_short Expression of nitric oxide synthases from photosynthetic microorganisms improves growth and stress tolerance in E. coli
title_full Expression of nitric oxide synthases from photosynthetic microorganisms improves growth and stress tolerance in E. coli
title_fullStr Expression of nitric oxide synthases from photosynthetic microorganisms improves growth and stress tolerance in E. coli
title_full_unstemmed Expression of nitric oxide synthases from photosynthetic microorganisms improves growth and stress tolerance in E. coli
title_sort Expression of nitric oxide synthases from photosynthetic microorganisms improves growth and stress tolerance in E. coli
dc.creator.none.fl_str_mv Correa Aragunde, Maria Natalia
Nejamkin, Andres
del Castello, Fiorella Paola
Foresi, Noelia Pamela
Lamattina, Lorenzo
author Correa Aragunde, Maria Natalia
author_facet Correa Aragunde, Maria Natalia
Nejamkin, Andres
del Castello, Fiorella Paola
Foresi, Noelia Pamela
Lamattina, Lorenzo
author_role author
author2 Nejamkin, Andres
del Castello, Fiorella Paola
Foresi, Noelia Pamela
Lamattina, Lorenzo
author2_role author
author
author
author
dc.subject.none.fl_str_mv BACTERIA
GROWTH
NITRIC OXIDE SYNTHASE
OXIDATIVE STRESS
topic BACTERIA
GROWTH
NITRIC OXIDE SYNTHASE
OXIDATIVE STRESS
purl_subject.fl_str_mv https://purl.org/becyt/ford/1.6
https://purl.org/becyt/ford/1
dc.description.none.fl_txt_mv Nitric oxide synthase (NOS) catalyzes the oxidation of the substrate L-Arginine (Arg) to produce citrulline and nitric oxide (NO). We have characterized NOS from two photosynthetic microorganisms: the NOS from the alga Ostreococcus tauri (OtNOS) and the cyanobacteria Synechococcus PCC 7335 (SyNOS). OtNOS and SyNOS possess distinct biochemical properties. OtNOS is a canonical NOS similar to animal NOS with an ultrafast NO producing activity. On the contrary, an extra globin domain present in SyNOS enzyme oxidizes over 70 % of the NO-produced to nitrate (NO3-). Here we describethe expression of recombinant OtNOS and SyNOS in Escherichia coli BL12 strain and analyze bacterial growth and tolerance to nitrosative stress. Results show that the E. coli cultures expressing OtNOS and SyNOS reach a higher OD at the exponential phase with respect to bacteria transformed with the empty vector (EV). This result correlates with higher NOS protein levels assayed by immunoblot, total protein and nitrate content in NOS recombinant strain cultures. Moreover, the expression of SyNOS and at less extent of OtNOS confers the ability to grow in minimal medium with Arg as a sole N source (and plenty C-source), suggesting that NOS enzymes are active in E. coli. The high NO producing activity reported in OtNOS correlates with the flavohemoglobin hmp induction in E. coli strain expressing OtNOS, suggesting that this strain senses nitrosative stress. Furthermore, nitrosative stress generated by the addition of 1 mM of the NO donor sodium nitroprusside (SNP) reduced growth rate (0.4-fold respect to no SNP addition) in bacterial culture expressing EV. However, the expression of recombinant OtNOS and at less extent SyNOS, attenuated SNP toxicity (0.8- and 0.6-fold, respectively, compared to no SNP addition). E. coli does not synthesize the major NOS cofactor tetrahydrobiopterin (BH4). Bioinformatics tools and ligand docking analysis were used to provide evidence supporting tetrahydromonapterin (MH4) as a possible pterin cofactor required for NOS catalytic activity in E. coli. These results open an exciting new window about the versatility of pterin cofactor working in the different NOSs dispersed in distant organisms along the life tree. In summary, our results show that NOS from photosynthetic microorganisms increases the growth and confers nitrosative stress tolerance in E. coli. Supported by AGENCIA, CONICET and UNMdP
Fil: Correa Aragunde, Maria Natalia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina
Fil: Nejamkin, Andres. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina
Fil: del Castello, Fiorella Paola. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina
Fil: Foresi, Noelia Pamela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina
Fil: Lamattina, Lorenzo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina
Reunión Conjunta SAIB-SAMIGE 2020. LVI Reunión Anual de la Sociedad Argentina de Investigación en Bioquímica y Biología Molecular. LVI Reunión Anual Sociedad Argentina de Bioquímica y Biología Molecular (SAIB). XV Reunión Anual Sociedad Argentina de Microbiología General (SAMIGE)
Buenos Aires
Argentina
Sociedad Argentina de Investigación en Bioquímica y Biología Molecular
Asociación Civil de Microbiología General
description Nitric oxide synthase (NOS) catalyzes the oxidation of the substrate L-Arginine (Arg) to produce citrulline and nitric oxide (NO). We have characterized NOS from two photosynthetic microorganisms: the NOS from the alga Ostreococcus tauri (OtNOS) and the cyanobacteria Synechococcus PCC 7335 (SyNOS). OtNOS and SyNOS possess distinct biochemical properties. OtNOS is a canonical NOS similar to animal NOS with an ultrafast NO producing activity. On the contrary, an extra globin domain present in SyNOS enzyme oxidizes over 70 % of the NO-produced to nitrate (NO3-). Here we describethe expression of recombinant OtNOS and SyNOS in Escherichia coli BL12 strain and analyze bacterial growth and tolerance to nitrosative stress. Results show that the E. coli cultures expressing OtNOS and SyNOS reach a higher OD at the exponential phase with respect to bacteria transformed with the empty vector (EV). This result correlates with higher NOS protein levels assayed by immunoblot, total protein and nitrate content in NOS recombinant strain cultures. Moreover, the expression of SyNOS and at less extent of OtNOS confers the ability to grow in minimal medium with Arg as a sole N source (and plenty C-source), suggesting that NOS enzymes are active in E. coli. The high NO producing activity reported in OtNOS correlates with the flavohemoglobin hmp induction in E. coli strain expressing OtNOS, suggesting that this strain senses nitrosative stress. Furthermore, nitrosative stress generated by the addition of 1 mM of the NO donor sodium nitroprusside (SNP) reduced growth rate (0.4-fold respect to no SNP addition) in bacterial culture expressing EV. However, the expression of recombinant OtNOS and at less extent SyNOS, attenuated SNP toxicity (0.8- and 0.6-fold, respectively, compared to no SNP addition). E. coli does not synthesize the major NOS cofactor tetrahydrobiopterin (BH4). Bioinformatics tools and ligand docking analysis were used to provide evidence supporting tetrahydromonapterin (MH4) as a possible pterin cofactor required for NOS catalytic activity in E. coli. These results open an exciting new window about the versatility of pterin cofactor working in the different NOSs dispersed in distant organisms along the life tree. In summary, our results show that NOS from photosynthetic microorganisms increases the growth and confers nitrosative stress tolerance in E. coli. Supported by AGENCIA, CONICET and UNMdP
publishDate 2020
dc.date.none.fl_str_mv 2020
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http://purl.org/coar/resource_type/c_5794
info:ar-repo/semantics/documentoDeConferencia
status_str publishedVersion
format conferenceObject
dc.identifier.none.fl_str_mv http://hdl.handle.net/11336/217194
Expression of nitric oxide synthases from photosynthetic microorganisms improves growth and stress tolerance in E. coli; Reunión Conjunta SAIB-SAMIGE 2020. LVI Reunión Anual de la Sociedad Argentina de Investigación en Bioquímica y Biología Molecular. LVI Reunión Anual Sociedad Argentina de Bioquímica y Biología Molecular (SAIB). XV Reunión Anual Sociedad Argentina de Microbiología General (SAMIGE); Buenos Aires; Argentina; 2020; 108-108
0327-9545
1667-5746
CONICET Digital
CONICET
url http://hdl.handle.net/11336/217194
identifier_str_mv Expression of nitric oxide synthases from photosynthetic microorganisms improves growth and stress tolerance in E. coli; Reunión Conjunta SAIB-SAMIGE 2020. LVI Reunión Anual de la Sociedad Argentina de Investigación en Bioquímica y Biología Molecular. LVI Reunión Anual Sociedad Argentina de Bioquímica y Biología Molecular (SAIB). XV Reunión Anual Sociedad Argentina de Microbiología General (SAMIGE); Buenos Aires; Argentina; 2020; 108-108
0327-9545
1667-5746
CONICET Digital
CONICET
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language eng
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