Imaging Ca2+ Concentration and pH in Nanopores/Channels of Protein Crystals.
Identifieur interne : 000053 ( Main/Curation ); précédent : 000052; suivant : 000054Imaging Ca2+ Concentration and pH in Nanopores/Channels of Protein Crystals.
Auteurs : Kazuo Mori [Japon] ; Bernd Kuhn [Japon]Source :
- The journal of physical chemistry. B [ 1520-5207 ] ; 2018.
Descripteurs français
- KwdFr :
- Calcium (composition chimique), Colorants fluorescents (composition chimique), Concentration en ions d'hydrogène (MeSH), Cristallisation (MeSH), Lysozyme (composition chimique), Mesures de luminescence (instrumentation), Mesures de luminescence (méthodes), Nanopores (MeSH), Protéines végétales (composition chimique).
- MESH :
- composition chimique : Calcium, Colorants fluorescents, Lysozyme, Protéines végétales.
- méthodes : Mesures de luminescence.
- instrumentation : Concentration en ions d'hydrogène, Cristallisation, Mesures de luminescence, Nanopores.
English descriptors
- KwdEn :
- MESH :
- chemical , chemistry : Calcium, Fluorescent Dyes, Muramidase, Plant Proteins.
- instrumentation : Luminescent Measurements.
- methods : Luminescent Measurements.
- Crystallization, Hydrogen-Ion Concentration, Nanopores.
Abstract
Protein crystals are nanoporous materials. Despite this important characteristic, little is known about the conditions in the pores, also called channels. Here, we describe a method to study the calcium concentration and pH in the nanopores of thaumatin and lysozyme crystals. We load the crystal nanopores with fluorescent indicators and then perfuse the crystals with solutions of different calcium concentrations and pH while reading out the crystal's fluorescence intensity with confocal microscopy. By calibrating the fluorescence signal, we can determine the calcium concentration and pH in the nanopores. For the pH in thaumatin nanopores measured with the ratiometric pH sensor SNARF-1, we find a -0.7 pH shift compared to the bath pH corresponding to a fivefold higher proton concentration. This is similar to the -0.3 pH shift found in lysozyme nanopores. With single-wavelength probes, we find that the calcium concentration in thaumatin crystal nanopores is the same as in the bath, whereas it is 0.24 times lower in lysozyme nanopores. Summarizing, our experiments show that calcium concentration and pH in the nanopores of protein crystals can deviate significantly from that in the bath. In general, the described method can be applied for testing a wide range of ion or small-molecule concentrations in transparent nanoporous materials not only with ratiometric but also with single wavelength fluorescent indicators.
DOI: 10.1021/acs.jpcb.8b07099
PubMed: 30351149
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<record><TEI><teiHeader><fileDesc><titleStmt><title xml:lang="en">Imaging Ca<sup>2+</sup> Concentration and pH in Nanopores/Channels of Protein Crystals.</title><author><name sortKey="Mori, Kazuo" sort="Mori, Kazuo" uniqKey="Mori K" first="Kazuo" last="Mori">Kazuo Mori</name><affiliation wicri:level="1"><nlm:affiliation>Okinawa Institute of Science and Technology , Graduate University , 1919-1 Tancha , Onna-son, Okinawa 904-0495 , Japan.</nlm:affiliation><country xml:lang="fr">Japon</country><wicri:regionArea>Okinawa Institute of Science and Technology , Graduate University , 1919-1 Tancha , Onna-son, Okinawa 904-0495 </wicri:regionArea></affiliation></author><author><name sortKey="Kuhn, Bernd" sort="Kuhn, Bernd" uniqKey="Kuhn B" first="Bernd" last="Kuhn">Bernd Kuhn</name><affiliation wicri:level="1"><nlm:affiliation>Okinawa Institute of Science and Technology , Graduate University , 1919-1 Tancha , Onna-son, Okinawa 904-0495 , Japan.</nlm:affiliation><country xml:lang="fr">Japon</country><wicri:regionArea>Okinawa Institute of Science and Technology , Graduate University , 1919-1 Tancha , Onna-son, Okinawa 904-0495 </wicri:regionArea></affiliation></author></titleStmt><publicationStmt><idno type="wicri:source">PubMed</idno><date when="2018">2018</date><idno type="RBID">pubmed:30351149</idno><idno type="pmid">30351149</idno><idno type="doi">10.1021/acs.jpcb.8b07099</idno><idno type="wicri:Area/Main/Corpus">000053</idno><idno type="wicri:explorRef" wicri:stream="Main" wicri:step="Corpus" wicri:corpus="PubMed">000053</idno><idno type="wicri:Area/Main/Curation">000053</idno><idno type="wicri:explorRef" wicri:stream="Main" wicri:step="Curation">000053</idno></publicationStmt><sourceDesc><biblStruct><analytic><title xml:lang="en">Imaging Ca<sup>2+</sup> Concentration and pH in Nanopores/Channels of Protein Crystals.</title><author><name sortKey="Mori, Kazuo" sort="Mori, Kazuo" uniqKey="Mori K" first="Kazuo" last="Mori">Kazuo Mori</name><affiliation wicri:level="1"><nlm:affiliation>Okinawa Institute of Science and Technology , Graduate University , 1919-1 Tancha , Onna-son, Okinawa 904-0495 , Japan.</nlm:affiliation><country xml:lang="fr">Japon</country><wicri:regionArea>Okinawa Institute of Science and Technology , Graduate University , 1919-1 Tancha , Onna-son, Okinawa 904-0495 </wicri:regionArea></affiliation></author><author><name sortKey="Kuhn, Bernd" sort="Kuhn, Bernd" uniqKey="Kuhn B" first="Bernd" last="Kuhn">Bernd Kuhn</name><affiliation wicri:level="1"><nlm:affiliation>Okinawa Institute of Science and Technology , Graduate University , 1919-1 Tancha , Onna-son, Okinawa 904-0495 , Japan.</nlm:affiliation><country xml:lang="fr">Japon</country><wicri:regionArea>Okinawa Institute of Science and Technology , Graduate University , 1919-1 Tancha , Onna-son, Okinawa 904-0495 </wicri:regionArea></affiliation></author></analytic><series><title level="j">The journal of physical chemistry. B</title><idno type="eISSN">1520-5207</idno><imprint><date when="2018" type="published">2018</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Calcium (chemistry)</term><term>Crystallization (MeSH)</term><term>Fluorescent Dyes (chemistry)</term><term>Hydrogen-Ion Concentration (MeSH)</term><term>Luminescent Measurements (instrumentation)</term><term>Luminescent Measurements (methods)</term><term>Muramidase (chemistry)</term><term>Nanopores (MeSH)</term><term>Plant Proteins (chemistry)</term></keywords><keywords scheme="KwdFr" xml:lang="fr"><term>Calcium (composition chimique)</term><term>Colorants fluorescents (composition chimique)</term><term>Concentration en ions d'hydrogène (MeSH)</term><term>Cristallisation (MeSH)</term><term>Lysozyme (composition chimique)</term><term>Mesures de luminescence (instrumentation)</term><term>Mesures de luminescence (méthodes)</term><term>Nanopores (MeSH)</term><term>Protéines végétales (composition chimique)</term></keywords><keywords scheme="MESH" type="chemical" qualifier="chemistry" xml:lang="en"><term>Calcium</term><term>Fluorescent Dyes</term><term>Muramidase</term><term>Plant Proteins</term></keywords><keywords scheme="MESH" qualifier="composition chimique" xml:lang="fr"><term>Calcium</term><term>Colorants fluorescents</term><term>Lysozyme</term><term>Protéines végétales</term></keywords><keywords scheme="MESH" qualifier="instrumentation" xml:lang="en"><term>Luminescent Measurements</term></keywords><keywords scheme="MESH" qualifier="methods" xml:lang="en"><term>Luminescent Measurements</term></keywords><keywords scheme="MESH" qualifier="méthodes" xml:lang="fr"><term>Mesures de luminescence</term></keywords><keywords scheme="MESH" xml:lang="en"><term>Crystallization</term><term>Hydrogen-Ion Concentration</term><term>Nanopores</term></keywords><keywords scheme="MESH" qualifier="instrumentation" xml:lang="fr"><term>Concentration en ions d'hydrogène</term><term>Cristallisation</term><term>Mesures de luminescence</term><term>Nanopores</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">Protein crystals are nanoporous materials. Despite this important characteristic, little is known about the conditions in the pores, also called channels. Here, we describe a method to study the calcium concentration and pH in the nanopores of thaumatin and lysozyme crystals. We load the crystal nanopores with fluorescent indicators and then perfuse the crystals with solutions of different calcium concentrations and pH while reading out the crystal's fluorescence intensity with confocal microscopy. By calibrating the fluorescence signal, we can determine the calcium concentration and pH in the nanopores. For the pH in thaumatin nanopores measured with the ratiometric pH sensor SNARF-1, we find a -0.7 pH shift compared to the bath pH corresponding to a fivefold higher proton concentration. This is similar to the -0.3 pH shift found in lysozyme nanopores. With single-wavelength probes, we find that the calcium concentration in thaumatin crystal nanopores is the same as in the bath, whereas it is 0.24 times lower in lysozyme nanopores. Summarizing, our experiments show that calcium concentration and pH in the nanopores of protein crystals can deviate significantly from that in the bath. In general, the described method can be applied for testing a wide range of ion or small-molecule concentrations in transparent nanoporous materials not only with ratiometric but also with single wavelength fluorescent indicators.</div></front></TEI><pubmed><MedlineCitation Status="MEDLINE" Owner="NLM"><PMID Version="1">30351149</PMID><DateCompleted><Year>2019</Year><Month>10</Month><Day>15</Day></DateCompleted><DateRevised><Year>2019</Year><Month>10</Month><Day>15</Day></DateRevised><Article PubModel="Print-Electronic"><Journal><ISSN IssnType="Electronic">1520-5207</ISSN><JournalIssue CitedMedium="Internet"><Volume>122</Volume><Issue>42</Issue><PubDate><Year>2018</Year><Month>10</Month><Day>25</Day></PubDate></JournalIssue><Title>The journal of physical chemistry. B</Title><ISOAbbreviation>J Phys Chem B</ISOAbbreviation></Journal><ArticleTitle>Imaging Ca<sup>2+</sup> Concentration and pH in Nanopores/Channels of Protein Crystals.</ArticleTitle><Pagination><MedlinePgn>9646-9653</MedlinePgn></Pagination><ELocationID EIdType="doi" ValidYN="Y">10.1021/acs.jpcb.8b07099</ELocationID><Abstract><AbstractText>Protein crystals are nanoporous materials. Despite this important characteristic, little is known about the conditions in the pores, also called channels. Here, we describe a method to study the calcium concentration and pH in the nanopores of thaumatin and lysozyme crystals. We load the crystal nanopores with fluorescent indicators and then perfuse the crystals with solutions of different calcium concentrations and pH while reading out the crystal's fluorescence intensity with confocal microscopy. By calibrating the fluorescence signal, we can determine the calcium concentration and pH in the nanopores. For the pH in thaumatin nanopores measured with the ratiometric pH sensor SNARF-1, we find a -0.7 pH shift compared to the bath pH corresponding to a fivefold higher proton concentration. This is similar to the -0.3 pH shift found in lysozyme nanopores. With single-wavelength probes, we find that the calcium concentration in thaumatin crystal nanopores is the same as in the bath, whereas it is 0.24 times lower in lysozyme nanopores. Summarizing, our experiments show that calcium concentration and pH in the nanopores of protein crystals can deviate significantly from that in the bath. In general, the described method can be applied for testing a wide range of ion or small-molecule concentrations in transparent nanoporous materials not only with ratiometric but also with single wavelength fluorescent indicators.</AbstractText></Abstract><AuthorList CompleteYN="Y"><Author ValidYN="Y"><LastName>Mori</LastName><ForeName>Kazuo</ForeName><Initials>K</Initials><AffiliationInfo><Affiliation>Okinawa Institute of Science and Technology , Graduate University , 1919-1 Tancha , Onna-son, Okinawa 904-0495 , Japan.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Kuhn</LastName><ForeName>Bernd</ForeName><Initials>B</Initials><Identifier Source="ORCID">0000-0002-6852-2433</Identifier><AffiliationInfo><Affiliation>Okinawa Institute of Science and Technology , Graduate University , 1919-1 Tancha , Onna-son, Okinawa 904-0495 , Japan.</Affiliation></AffiliationInfo></Author></AuthorList><Language>eng</Language><PublicationTypeList><PublicationType UI="D016428">Journal Article</PublicationType><PublicationType UI="D013485">Research Support, Non-U.S. Gov't</PublicationType></PublicationTypeList><ArticleDate DateType="Electronic"><Year>2018</Year><Month>10</Month><Day>11</Day></ArticleDate></Article><MedlineJournalInfo><Country>United States</Country><MedlineTA>J Phys Chem B</MedlineTA><NlmUniqueID>101157530</NlmUniqueID><ISSNLinking>1520-5207</ISSNLinking></MedlineJournalInfo><ChemicalList><Chemical><RegistryNumber>0</RegistryNumber><NameOfSubstance UI="D005456">Fluorescent Dyes</NameOfSubstance></Chemical><Chemical><RegistryNumber>0</RegistryNumber><NameOfSubstance UI="D010940">Plant Proteins</NameOfSubstance></Chemical><Chemical><RegistryNumber>53850-34-3</RegistryNumber><NameOfSubstance UI="C003427">thaumatin protein, plant</NameOfSubstance></Chemical><Chemical><RegistryNumber>EC 3.2.1.17</RegistryNumber><NameOfSubstance UI="D009113">Muramidase</NameOfSubstance></Chemical><Chemical><RegistryNumber>SY7Q814VUP</RegistryNumber><NameOfSubstance UI="D002118">Calcium</NameOfSubstance></Chemical></ChemicalList><CitationSubset>IM</CitationSubset><MeshHeadingList><MeshHeading><DescriptorName UI="D002118" MajorTopicYN="N">Calcium</DescriptorName><QualifierName UI="Q000737" MajorTopicYN="Y">chemistry</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D003460" MajorTopicYN="N">Crystallization</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D005456" MajorTopicYN="N">Fluorescent Dyes</DescriptorName><QualifierName UI="Q000737" MajorTopicYN="N">chemistry</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D006863" MajorTopicYN="N">Hydrogen-Ion Concentration</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D008163" MajorTopicYN="N">Luminescent Measurements</DescriptorName><QualifierName UI="Q000295" MajorTopicYN="N">instrumentation</QualifierName><QualifierName UI="Q000379" MajorTopicYN="N">methods</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D009113" MajorTopicYN="N">Muramidase</DescriptorName><QualifierName UI="Q000737" MajorTopicYN="Y">chemistry</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D058608" MajorTopicYN="N">Nanopores</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D010940" MajorTopicYN="N">Plant Proteins</DescriptorName><QualifierName UI="Q000737" MajorTopicYN="Y">chemistry</QualifierName></MeshHeading></MeshHeadingList></MedlineCitation><PubmedData><History><PubMedPubDate PubStatus="pubmed"><Year>2018</Year><Month>10</Month><Day>24</Day><Hour>6</Hour><Minute>0</Minute></PubMedPubDate><PubMedPubDate PubStatus="medline"><Year>2019</Year><Month>10</Month><Day>16</Day><Hour>6</Hour><Minute>0</Minute></PubMedPubDate><PubMedPubDate PubStatus="entrez"><Year>2018</Year><Month>10</Month><Day>24</Day><Hour>6</Hour><Minute>0</Minute></PubMedPubDate></History><PublicationStatus>ppublish</PublicationStatus><ArticleIdList><ArticleId IdType="pubmed">30351149</ArticleId><ArticleId IdType="doi">10.1021/acs.jpcb.8b07099</ArticleId></ArticleIdList></PubmedData></pubmed></record>Pour manipuler ce document sous Unix (Dilib)
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