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Comparison of the Catalytic Activities of Three Isozymes of Carnitine Palmitoyltransferase 1 Expressed in COS7 Cells

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Abstract

The enzyme carnitine palmitoyltransferase 1 (CPT1) catalyzes the transfer of an acyl group from acyl-CoA to carnitine to form acylcarnitine, and three isozymes of it, 1a, 1b, and 1c, have been identified. Interestingly, the 1c isozyme was reported to show no enzymatic activity, but it was not clearly demonstrated whether this inactivity was due to its dysfunction or due to its poor expression. In the present study, we (a) expressed individual CPT1 isozymes in COS7 cells, (b) evaluated quantitatively their expression levels by Western blotting using the three bacterially expressed CPT1 isozymes as standards, and (c) evaluated their catalytic activities. With these experiments, we successfully demonstrated that the absence of the enzymatic activity of the 1c isozyme was due to its dysfunction. In addition, experiments on the preparation of standard CPT1 isozymes revealed that the 1c isozyme did not show the standard relationship between migration in an SDS–PAGE gel and molecular size. We further tried to determine why the 1c isozyme was inert by preparing chimeric CPT1 between 1a and 1c, but no clear conclusion could be drawn because one of the chimeric CPT1s was not sufficiently expressed.

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Abbreviations

CPT1:

Carnitine palmitoyltransferase 1

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Acknowledgments

This study was supported by the Program for the Promotion of Basic and Applied Researches for Innovations in Bio-oriented Industry (BRAIN).

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Authors and Affiliations

  1. Institute for Genome Research, University of Tokushima, Kuramoto-cho-3, Tokushima, 770-8503, Japan

    Takuya Hada, Takenori Yamamoto, Atsushi Yamamoto & Yasuo Shinohara

  2. Graduate School of Pharmaceutical Sciences, University of Tokushima, Shomachi-1, Tokushima, 770-8505, Japan

    Takuya Hada, Takenori Yamamoto, Atsushi Yamamoto, Naoshi Yamazaki, Yoshiharu Takiguchi & Yasuo Shinohara

  3. Faculty of Pharmaceutical Science, Suzuka University of Medical Science, Minamitamagaki-cho-3500, Suzuka, 513-8670, Japan

    Kazuto Ohkura

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  1. Takuya Hada
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  2. Takenori Yamamoto
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Correspondence to Yasuo Shinohara.

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Hada, T., Yamamoto, T., Yamamoto, A. et al. Comparison of the Catalytic Activities of Three Isozymes of Carnitine Palmitoyltransferase 1 Expressed in COS7 Cells. Appl Biochem Biotechnol 172, 1486–1496 (2014). https://doi.org/10.1007/s12010-013-0619-y

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