Tissue nonspecific alkaline phosphatase is activated via a two-step mechanism by zinc transport complexes in the early secretory pathway

Ayako Fukunaka, Yayoi Kurokawa, Fumie Teranishi, Israel Sekler, Kimimitsu Oda, M. Leigh Ackland, Victor Faundez, Makoto Hiromura, Seiji Masuda, Masaya Nagao, Shuichi Enomoto, Taiho Kambe

Research output: Contribution to journalArticle

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Abstract

A number of enzymes become functional by binding to zinc during their journey through the early secretory pathway. The zinc transporters (ZnTs) located there play important roles in this step. We have previously shown that two zinc transport complexes, ZnT5/ZnT6 heterodimers and ZnT7 homo-oligomers, are required for the activation of alkaline phosphatases, by converting them from the apo- to the holo-form. Here, we investigated the molecular mechanisms of this activation. ZnT1 and ZnT4 expressed in chicken DT40 cells did not contribute to the activation of tissue nonspecific alkaline phosphatase (TNAP). The reduced activity of TNAP in DT40 cells deficient in both ZnT complexes was not restored by zinc supplementation nor by exogenous expression of other ZnTs that increase the zinc content in the secretory pathway. Moreover, we showed that expression of ZnT5/ZnT6 heterodimers reconstituted with zinc transport-incompetent ZnT5 mutant failed to restore TNAP activity but could stabilize the TNAP protein as the apo-form, regardless of zinc status. These findings demonstrate that TNAP is activated not simply by passive zinc binding but by an elaborate two-step mechanism via protein stabilization followed by enzyme conversion from the apo- to the holo-form with zinc loaded by ZnT complexes in the early secretory pathway.

Original languageEnglish
Pages (from-to)16363-16373
Number of pages11
JournalJournal of Biological Chemistry
Volume286
Issue number18
DOIs
Publication statusPublished - May 6 2011

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Secretory Pathway
Alkaline Phosphatase
Zinc
Tissue
Chemical activation
Enzymes
Oligomers
Chickens
Proteins
Stabilization

ASJC Scopus subject areas

  • Biochemistry
  • Cell Biology
  • Molecular Biology

Cite this

Tissue nonspecific alkaline phosphatase is activated via a two-step mechanism by zinc transport complexes in the early secretory pathway. / Fukunaka, Ayako; Kurokawa, Yayoi; Teranishi, Fumie; Sekler, Israel; Oda, Kimimitsu; Ackland, M. Leigh; Faundez, Victor; Hiromura, Makoto; Masuda, Seiji; Nagao, Masaya; Enomoto, Shuichi; Kambe, Taiho.

In: Journal of Biological Chemistry, Vol. 286, No. 18, 06.05.2011, p. 16363-16373.

Research output: Contribution to journalArticle

Fukunaka, A, Kurokawa, Y, Teranishi, F, Sekler, I, Oda, K, Ackland, ML, Faundez, V, Hiromura, M, Masuda, S, Nagao, M, Enomoto, S & Kambe, T 2011, 'Tissue nonspecific alkaline phosphatase is activated via a two-step mechanism by zinc transport complexes in the early secretory pathway', Journal of Biological Chemistry, vol. 286, no. 18, pp. 16363-16373. https://doi.org/10.1074/jbc.M111.227173
Fukunaka, Ayako ; Kurokawa, Yayoi ; Teranishi, Fumie ; Sekler, Israel ; Oda, Kimimitsu ; Ackland, M. Leigh ; Faundez, Victor ; Hiromura, Makoto ; Masuda, Seiji ; Nagao, Masaya ; Enomoto, Shuichi ; Kambe, Taiho. / Tissue nonspecific alkaline phosphatase is activated via a two-step mechanism by zinc transport complexes in the early secretory pathway. In: Journal of Biological Chemistry. 2011 ; Vol. 286, No. 18. pp. 16363-16373.
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