Biotin Carboxyl Carrier Protein and Carboxyltransferase Subunits of the Multi-subunit Form of acetyl-CoA carboxylase from Brassica napus: Cloning and Analysis of Expression during Oilseed Rape Embryogenesis

Authors

Kieran M. Elborough, University of Durham
Robert Winz, University of Durham
Ranjit K. Deka, Brassica and Oilseeds Research Department, John Innes Institute, Norwich
Jennifer E. Markham, University of Nebraska-LincolnFollow
Andrew J. White, University of Durham
Stephen Rawsthorne, Brassica and Oilseeds Research Department, John Innes Institute, Norwich
Antoni R. Slabas, University of Durham

ORCID IDs

0000-0003-2397-3131 (JEM)

Document Type

Article

Date of this Version

4-1-1996

Citation

Biochemical Journal (April 1, 1996) 315 (Part 1): 103-112

doi: 10.1042/bj3150103

Comments

Copyright 1996, Biochemical Society/Portland Press. Used by permission.

Abstract

In the oilseed rape Brassica napus there are two forms of acetyl- CoA carboxylase (ACCase). As in other dicotyledonous plants there is a type I ACCase, the single polypeptide 220 kDa form, and a type II multi-subunit complex analogous to that of Escherichia coli and Anabaena. This paper describes the cloning and characterization of a plant biotin carboxyl carrier protein (BCCP) from the type II ACCase complex that shows 61% identity/79% similarity with Anabaena BCCP at the amino acid level. Six classes of nuclear encoded oilseed rape BCCP cDNA were cloned, two of which contained the entire coding region. The BCCP sequences allowed the assignment of function to two previously unassigned Arabidopsis expressed sequence tag (EST) sequences. We also report the cloning of a second type II ACCase component from oilseed rape, the β-carboxyltransferase subunit (βCT), which is chloroplast-encoded. Northern analysis showed that although the relative levels of BCCP and βCT mRNA differed between different oilseed rape tissues, their temporal patterns of expression were identical during embryo development. At the protein level, expression of BCCP during embryo development was studied by Western blotting, using affinity-purified anti-biotin polyclonal sera. With this technique a 35 kDa protein thought to be BCCP was shown to reside within the chloroplast. This analysis also permitted us to view the differential expression of several unidentified biotinylated proteins during embryogenesis.