Neurochemical Research

, Volume 36, Issue 8, pp 1426–1434 | Cite as

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

  • Ioanna Chalatsa
  • Emmanuel G. Fragoulis
  • Dido Vassilacopoulou
Original Paper

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.

Keywords

L-Dopa decarboxylase Membrane fraction Protease inhibitors Solubilization Triton X-114 

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

Notes

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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Copyright information

© Springer Science+Business Media, LLC 2011

Authors and Affiliations

  • Ioanna Chalatsa
    • 1
  • Emmanuel G. Fragoulis
    • 1
  • Dido Vassilacopoulou
    • 1
  1. 1.Department of Biochemistry and Molecular BiologyUniversity of AthensAthensGreece

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