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Release of Membrane-Associated L-Dopa Decarboxylase from Human Cells

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Abstract

L-Dopa Decarboxylase is a pyridoxal 5-phosphate (PLP)-dependent enzyme that catalyses the decarboxylation of L-Dopa to dopamine. In this study, we investigated the cellular topology of the active human enzyme. Fractionation of membranes from human cell lines, of neural and non-neural origin, by temperature-induced phase separation in Triton X-114 resulted in the detection of DDC molecules in all separation phases. Solubilization of membrane-associated DDC was observed in a pH and time-dependent manner and was affected by divalent cations and protease inhibitors, suggesting the involvement of a possible release mechanism. The study of the biological properties and function of the solubilization phenomenon described here, as well as, the study of the membrane-associated enzyme could provide us with new information about the participation of the human L-Dopa decarboxylase in physiological and aberrant processes.

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Abbreviations

DDC:

L-Dopa decarboxylase

PLP:

Pyridoxal 5-phosphate

HEK-293:

Human embryonic kidney

SH-SY5Y:

Human neuroblastoma

HTB-14:

Human glioblastoma, astrocytoma

NEM:

N-ethylmaleimide

SCLC:

Small cell lung carcinomas

DBH:

Dopamine β-hydroxylase

TH:

Tyrosine hydroxylase

PNMT:

Phenylethanolamine N-methyl Transferase

G-PI-anchor:

Glycosyl-phosphatidylinositol-anchor

MF:

Membrane fraction

APS I:

Autoimmune polyendocrine syndrome type I

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Aknowledgments

This work represents part of the Doctoral thesis of Ms Ioanna Chalatsa. We would like to thank Ms S. Tsiropoulou for her assistance.

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Authors and Affiliations

  1. Department of Biochemistry and Molecular Biology, University of Athens, Panepistimiopolis, Zografou, 15701, Athens, Greece

    Ioanna Chalatsa, Emmanuel G. Fragoulis & Dido Vassilacopoulou

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  1. Ioanna Chalatsa
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  2. Emmanuel G. Fragoulis
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  3. Dido Vassilacopoulou
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Correspondence to Dido Vassilacopoulou.

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Chalatsa, I., Fragoulis, E.G. & Vassilacopoulou, D. Release of Membrane-Associated L-Dopa Decarboxylase from Human Cells. Neurochem Res 36, 1426–1434 (2011). https://doi.org/10.1007/s11064-011-0468-4

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