Resonance Raman spectroscopic investigation of the light-harvesting chromophore in Escherichia coli photolyase and Vibrio cholerae cryptochrome-1

Olga Sokolova, Christine Cecala, Anand Gopal, Frank Cortazar, Carla McDowell-Buchanan, Aziz Sancar, Yvonne Gindt, Johannes Schelvis

Research output: Contribution to journalArticle

9 Citations (Scopus)

Abstract

Photolyases and cryptochromes are flavoproteins that belong to the class of blue-light photoreceptors. They usually bind two chromophores: flavin adenine dinucleotide (FAD), which forms the active site, and a light-harvesting pigment, which is a 5,10-methenyltetrahydrofolate polyglutamate (MTHF) in most cases. In Escherichia coli photolyase (EcPhr), the MTHF cofactor is present in substoichiometric amounts after purification, while in Vibrio cholerae cryptochrome-1 (VcCry1) the MTHF cofactor is bound more strongly and is present at stoichiometric levels after purification. In this paper, we have used resonance Raman spectroscopy to monitor the effect of loss of MTHF on the protein-FAD interactions in EcPhr and to probe the protein-MTHF interactions in both EcPhr and VcCry1. We find that removal of MTHF does not perturb protein-FAD interactions, suggesting that it may not affect the physicochemical properties of FAD in EcPhr. Our data demonstrate that the pteridine ring of MTHF in EcPhr has different interactions with the protein matrix than that of MTHF in VcCry1. Comparison to solution resonance Raman spectra of MTHF suggests that the carbonyl of its pteridine ring in EcPhr experiences stronger hydrogen bonding and a more polar environment than in VcCry1, but that hydrogen bonding to the pteridine ring amine hydrogens is stronger in VcCry-1. These differences in hydrogen bonding may account for the higher binding affinity of MTHF in VcCry1 compared to EcPhr.

Original languageEnglish
Pages (from-to)3673-3681
Number of pages9
JournalBiochemistry
Volume46
Issue number12
DOIs
StatePublished - 27 Mar 2007

Fingerprint

Cryptochromes
Deoxyribodipyrimidine Photo-Lyase
Polyglutamic Acid
Vibrio cholerae
Chromophores
Escherichia coli
Light
Flavin-Adenine Dinucleotide
Pteridines
Hydrogen Bonding
Hydrogen bonds
Purification
Proteins
5,10-methenyltetrahydrofolate
Flavoproteins
Raman Spectrum Analysis
Pigments
Amines
Raman spectroscopy
Raman scattering

Cite this

Sokolova, Olga ; Cecala, Christine ; Gopal, Anand ; Cortazar, Frank ; McDowell-Buchanan, Carla ; Sancar, Aziz ; Gindt, Yvonne ; Schelvis, Johannes. / Resonance Raman spectroscopic investigation of the light-harvesting chromophore in Escherichia coli photolyase and Vibrio cholerae cryptochrome-1. In: Biochemistry. 2007 ; Vol. 46, No. 12. pp. 3673-3681.
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abstract = "Photolyases and cryptochromes are flavoproteins that belong to the class of blue-light photoreceptors. They usually bind two chromophores: flavin adenine dinucleotide (FAD), which forms the active site, and a light-harvesting pigment, which is a 5,10-methenyltetrahydrofolate polyglutamate (MTHF) in most cases. In Escherichia coli photolyase (EcPhr), the MTHF cofactor is present in substoichiometric amounts after purification, while in Vibrio cholerae cryptochrome-1 (VcCry1) the MTHF cofactor is bound more strongly and is present at stoichiometric levels after purification. In this paper, we have used resonance Raman spectroscopy to monitor the effect of loss of MTHF on the protein-FAD interactions in EcPhr and to probe the protein-MTHF interactions in both EcPhr and VcCry1. We find that removal of MTHF does not perturb protein-FAD interactions, suggesting that it may not affect the physicochemical properties of FAD in EcPhr. Our data demonstrate that the pteridine ring of MTHF in EcPhr has different interactions with the protein matrix than that of MTHF in VcCry1. Comparison to solution resonance Raman spectra of MTHF suggests that the carbonyl of its pteridine ring in EcPhr experiences stronger hydrogen bonding and a more polar environment than in VcCry1, but that hydrogen bonding to the pteridine ring amine hydrogens is stronger in VcCry-1. These differences in hydrogen bonding may account for the higher binding affinity of MTHF in VcCry1 compared to EcPhr.",
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Resonance Raman spectroscopic investigation of the light-harvesting chromophore in Escherichia coli photolyase and Vibrio cholerae cryptochrome-1. / Sokolova, Olga; Cecala, Christine; Gopal, Anand; Cortazar, Frank; McDowell-Buchanan, Carla; Sancar, Aziz; Gindt, Yvonne; Schelvis, Johannes.

In: Biochemistry, Vol. 46, No. 12, 27.03.2007, p. 3673-3681.

Research output: Contribution to journalArticle

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T1 - Resonance Raman spectroscopic investigation of the light-harvesting chromophore in Escherichia coli photolyase and Vibrio cholerae cryptochrome-1

AU - Sokolova, Olga

AU - Cecala, Christine

AU - Gopal, Anand

AU - Cortazar, Frank

AU - McDowell-Buchanan, Carla

AU - Sancar, Aziz

AU - Gindt, Yvonne

AU - Schelvis, Johannes

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AB - Photolyases and cryptochromes are flavoproteins that belong to the class of blue-light photoreceptors. They usually bind two chromophores: flavin adenine dinucleotide (FAD), which forms the active site, and a light-harvesting pigment, which is a 5,10-methenyltetrahydrofolate polyglutamate (MTHF) in most cases. In Escherichia coli photolyase (EcPhr), the MTHF cofactor is present in substoichiometric amounts after purification, while in Vibrio cholerae cryptochrome-1 (VcCry1) the MTHF cofactor is bound more strongly and is present at stoichiometric levels after purification. In this paper, we have used resonance Raman spectroscopy to monitor the effect of loss of MTHF on the protein-FAD interactions in EcPhr and to probe the protein-MTHF interactions in both EcPhr and VcCry1. We find that removal of MTHF does not perturb protein-FAD interactions, suggesting that it may not affect the physicochemical properties of FAD in EcPhr. Our data demonstrate that the pteridine ring of MTHF in EcPhr has different interactions with the protein matrix than that of MTHF in VcCry1. Comparison to solution resonance Raman spectra of MTHF suggests that the carbonyl of its pteridine ring in EcPhr experiences stronger hydrogen bonding and a more polar environment than in VcCry1, but that hydrogen bonding to the pteridine ring amine hydrogens is stronger in VcCry-1. These differences in hydrogen bonding may account for the higher binding affinity of MTHF in VcCry1 compared to EcPhr.

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