Protein gene product 9.5 (PGP9.5) is a cytosolic protein that is highly expressed in vertebrate neurons, which is now included in the ubiquitin C-terminal hydrolase subclass (UCH) on the basis of primary-structure homology and hydrolytic activity on the synthetic substrate ubiquitin ethyl ester (UbOEt). Some UCHs show affinity for immobilized ubiquitin, a property exploited to purify them. In this study we show that this property can also be applied to PGP9.5, since a protein has been purified to homogeneity from bovine retina by affinity chromatography on a ubiquitin–Sepharose column that can be identified with: (a) PGP9.5 with respect to molecular mass, primary structure and immunological reactivity; (b) the known UCHs with respect to some catalytic properties, such as hydrolytic activity on UbOEt, (which also characterizes PGP9.5), Km value and reactivity with cysteine and histidine-specific reagents. However, it differs with respect to other properties, e.g. inhibition by UbOEt and a wider pH range of activity.
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September 1996
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Research Article|
September 01 1996
Affinity purification and characterization of protein gene product 9.5 (PGP9.5) from retina
Marco PICCININI;
Marco PICCININI
*Dipartimento di Morfofisiologia Veterinaria, Università di Torino, Via Nizza 52, I-10126 Turin, Italy
†Dipartimento di Medicina e Oncologia Sperimentale, sezione di Biochimica, Università di Torino, Via Michelangelo, 27, I-10126 Turin, Italy
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Adalberto MERIGHI;
Adalberto MERIGHI
*Dipartimento di Morfofisiologia Veterinaria, Università di Torino, Via Nizza 52, I-10126 Turin, Italy
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Renato BRUNO;
Renato BRUNO
*Dipartimento di Morfofisiologia Veterinaria, Università di Torino, Via Nizza 52, I-10126 Turin, Italy
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Paolo CASCIO;
Paolo CASCIO
*Dipartimento di Morfofisiologia Veterinaria, Università di Torino, Via Nizza 52, I-10126 Turin, Italy
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Magda CURTO;
Magda CURTO
†Dipartimento di Medicina e Oncologia Sperimentale, sezione di Biochimica, Università di Torino, Via Michelangelo, 27, I-10126 Turin, Italy
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Silvia Clarissa MIOLETTI;
Silvia Clarissa MIOLETTI
†Dipartimento di Medicina e Oncologia Sperimentale, sezione di Biochimica, Università di Torino, Via Michelangelo, 27, I-10126 Turin, Italy
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Maria T. RINAUDO
Maria T. RINAUDO
‡
†Dipartimento di Medicina e Oncologia Sperimentale, sezione di Biochimica, Università di Torino, Via Michelangelo, 27, I-10126 Turin, Italy
‡To whom correspondence should be addressed.
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Publisher: Portland Press Ltd
Received:
January 24 1996
Revision Received:
March 25 1996
Accepted:
May 21 1996
Online ISSN: 1470-8728
Print ISSN: 0264-6021
The Biochemical Society, London © 1996
1996
Biochem J (1996) 318 (2): 711–716.
Article history
Received:
January 24 1996
Revision Received:
March 25 1996
Accepted:
May 21 1996
Citation
Marco PICCININI, Adalberto MERIGHI, Renato BRUNO, Paolo CASCIO, Magda CURTO, Silvia Clarissa MIOLETTI, Maria T. RINAUDO; Affinity purification and characterization of protein gene product 9.5 (PGP9.5) from retina. Biochem J 1 September 1996; 318 (2): 711–716. doi: https://doi.org/10.1042/bj3180711
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