Invariant Ser211 is involved in the catalysis of PD-L4, type I RIP from Phytolacca dioica leaves (570 views)
Proteins (ISSN: 0887-3585, 1097-0134, 1097-0134electronic), 2007 Apr 1; 67(1): 209-218.
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Paper type: Journal Article,
Impact factor: 3.354, 5-year impact factor: 3.905
Url: http://www.scopus.com/inward/record.url?eid=2-s2.0-33847399669&partnerID=40&md5=488bd05d9de230aaac9d56c1395edf1d
Keywords: 3d Modelling, Adenine Polynucleotide Glycosylases, Circular Dichroic Spectroscopy, Docking Analysis, Fluorescence Analyses, Ribosome Inactivating Proteins, Complementary Dna, Dna Glycosyltransferase, Polyadenylic Acid, Ribosome Rna, Serine, Unclassified Drug, Article, Catalysis, Enzyme Activity, Enzyme Substrate, Fluorescence Spectroscopy, Molecular Model, Nonhuman, Plant Leaf, Pokeweed, Priority Journal, Protein Purification, Three Dimensional Imaging, Amino Acid Sequence, Animals, Cell-Free System, Cloning, Computer Simulation, Escherichia Coli, Molecular Sequence Data, Mutagenesis, Site-Directed, N-Glycosyl Hydrolases, Phytolacca, Plant Leaves, Plant Proteins, Rabbits, Reticulocytes, Sequence Alignment, Spectrometry, Phytolacca Dioica,
Affiliations:
*** IBB - CNR ***
Dipartimento di Scienze della Vita, Seconda Università di Napoli, Via Vivaldi 43, I-81100 Caserta, Italy
Istituto di Biostrutture e Bioimmagini, CNR, via Mezzocannone 16, I-80138 Napoli, Italy
Dipartimento di Biochimica e Biofisica, CRISCEB (Centro di Ricerca Interdipartimentale per le Scienze Computazionali e Biotecnologiche), Seconda Università di Napoli, Via Costantinopoli 16, I-80138 Napoli, Italy
Istituto di Scienze dell'Alimentazione, CNR, Via Roma 52 A/C, 83100 Avellino, Italy
References:
Not available.
Proteins (ISSN: 0887-3585, 1097-0134, 1097-0134electronic), 2007 Apr 1; 67(1): 209-218.
Export to BibTeX Export to EndNote
Paper type: Journal Article,
Impact factor: 3.354, 5-year impact factor: 3.905
Url: http://www.scopus.com/inward/record.url?eid=2-s2.0-33847399669&partnerID=40&md5=488bd05d9de230aaac9d56c1395edf1d
Keywords: 3d Modelling, Adenine Polynucleotide Glycosylases, Circular Dichroic Spectroscopy, Docking Analysis, Fluorescence Analyses, Ribosome Inactivating Proteins, Complementary Dna, Dna Glycosyltransferase, Polyadenylic Acid, Ribosome Rna, Serine, Unclassified Drug, Article, Catalysis, Enzyme Activity, Enzyme Substrate, Fluorescence Spectroscopy, Molecular Model, Nonhuman, Plant Leaf, Pokeweed, Priority Journal, Protein Purification, Three Dimensional Imaging, Amino Acid Sequence, Animals, Cell-Free System, Cloning, Computer Simulation, Escherichia Coli, Molecular Sequence Data, Mutagenesis, Site-Directed, N-Glycosyl Hydrolases, Phytolacca, Plant Leaves, Plant Proteins, Rabbits, Reticulocytes, Sequence Alignment, Spectrometry, Phytolacca Dioica,
Affiliations:
*** IBB - CNR ***
Dipartimento di Scienze della Vita, Seconda Università di Napoli, Via Vivaldi 43, I-81100 Caserta, Italy
Istituto di Biostrutture e Bioimmagini, CNR, via Mezzocannone 16, I-80138 Napoli, Italy
Dipartimento di Biochimica e Biofisica, CRISCEB (Centro di Ricerca Interdipartimentale per le Scienze Computazionali e Biotecnologiche), Seconda Università di Napoli, Via Costantinopoli 16, I-80138 Napoli, Italy
Istituto di Scienze dell'Alimentazione, CNR, Via Roma 52 A/C, 83100 Avellino, Italy
References:
Not available.
Invariant Ser211 is involved in the catalysis of PD-L4, type I RIP from Phytolacca dioica leaves
Multiple sequence alignment analysis of ribosome inactivating proteins (RIPs) has revealed the occurrence of an invariant seryl residue in proximity of the catalytic tryptophan. The involvement of this seryl residue in the catalytic mechanism of RIPs was investigated by site-directed mutagenesis in PD-L4, type 1 RIP isolated from Phytolacca dioica leaves. We show that the replacement of Ser211 with Ala apparently does not influence the N-β-glycosidase activity on ribosomes (determined as IC50 in a cell-free system), but it reduces the adenine polynucleotide glycosylase activity (APG), assayed spectrophotometrically on other substrates such as DNA, rRNA, and poly(A). The ability of PDL4 to deadenylate polynucleotides appears more sensitive to the Ser211Ala replacement when poly(A) is used as substrate, as only 33% activity is retained by the mutant, while with more complex and heterogeneous substrates such as DNA and rRNA, its APG activity is 73% and 66%, respectively. While the mutated protein shows a conserved secondary structure by CD, it also exhibits a remarkably enhanced tryptophan fluorescence. This indicates that, although the overall protein tridimensional structure is maintained, removal of the hydrosyl group locally affects the environment of a Trp residue. Modelling and docking analyses confirm the interaction between Ser211 and Trp207, which is located within the active site, thus affecting RIP adenine polynucleotide glycosylase activity. Data accumulated so far confirm the potential involvement of Ser211 in the catalytic mechanism of type 1 RIP PD-L4 and a possible role in stabilizing the conformation of Trp207 side chain, which participates actively in the protein enzymatic activity. © 2007 Wiley-Liss, Inc.
Multiple sequence alignment analysis of ribosome inactivating proteins (RIPs) has revealed the occurrence of an invariant seryl residue in proximity of the catalytic tryptophan. The involvement of this seryl residue in the catalytic mechanism of RIPs was investigated by site-directed mutagenesis in PD-L4, type 1 RIP isolated from Phytolacca dioica leaves. We show that the replacement of Ser211 with Ala apparently does not influence the N-β-glycosidase activity on ribosomes (determined as IC50 in a cell-free system), but it reduces the adenine polynucleotide glycosylase activity (APG), assayed spectrophotometrically on other substrates such as DNA, rRNA, and poly(A). The ability of PDL4 to deadenylate polynucleotides appears more sensitive to the Ser211Ala replacement when poly(A) is used as substrate, as only 33% activity is retained by the mutant, while with more complex and heterogeneous substrates such as DNA and rRNA, its APG activity is 73% and 66%, respectively. While the mutated protein shows a conserved secondary structure by CD, it also exhibits a remarkably enhanced tryptophan fluorescence. This indicates that, although the overall protein tridimensional structure is maintained, removal of the hydrosyl group locally affects the environment of a Trp residue. Modelling and docking analyses confirm the interaction between Ser211 and Trp207, which is located within the active site, thus affecting RIP adenine polynucleotide glycosylase activity. Data accumulated so far confirm the potential involvement of Ser211 in the catalytic mechanism of type 1 RIP PD-L4 and a possible role in stabilizing the conformation of Trp207 side chain, which participates actively in the protein enzymatic activity. © 2007 Wiley-Liss, Inc.
Invariant Ser211 is involved in the catalysis of PD-L4, type I RIP from Phytolacca dioica leaves
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Invariant Ser211 is involved in the catalysis of PD-L4, type I RIP from Phytolacca dioica leaves
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Di Maro A, Berisio R, Ruggiero A, Tamburino R, Severino V, Zacchia E, Parente A
* Structural and enzymatic properties of an in vivo proteolytic form of PD-S2, type 1 ribosome-inactivating protein from seeds of Phytolacca dioica (796 views)
Biochem Biophys Res Commun (ISSN: 0006-291x), 2012 May 11; 421(3): 514-520.
Impact Factor: 2.406
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* Structural and enzymatic properties of an in vivo proteolytic form of PD-S2, type 1 ribosome-inactivating protein from seeds of Phytolacca dioica (796 views)
Biochem Biophys Res Commun (ISSN: 0006-291x), 2012 May 11; 421(3): 514-520.
Impact Factor: 2.406
View Export to BibTeX Export to EndNote
1 Records (1 excluding Abstracts).
Total impact factor: 2.406 (2.406 excluding Abstracts).
Total 5 year impact factor: 2.5 (2.5 excluding Abstracts).
Total impact factor: 2.406 (2.406 excluding Abstracts).
Total 5 year impact factor: 2.5 (2.5 excluding Abstracts).
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