Original Article

Mutational analysis of apolipoprotein B mRNA editing enzyme (APOBEC1): structure–function relationships of RNA editing and dimerization

Correspondence to: Ba-Bie Teng

APOBEC1 is the catalytic subunit of an enzyme complex that mediates apolipoprotein (apo) B mRNA editing. It dimerizes in vitro and requires complementation factor(s) for its editing activity. We have performed a systematic analysis of the structure–functional relationship of APOBEC1 by targeted mutagenesis of various sequence motifs within the protein. Using in vitro RNA editing assay, we found that basic amino acid clusters at the amino-terminal region R¹⁵R¹⁶R¹⁷ and R³³K³⁴, are essential for apoB mRNA editing. Mutation of R¹⁵R¹⁶R¹⁷ to K¹⁵K¹⁶K¹⁷ and mutation of R³³K³⁴ simultaneously to A³³A³⁴ almost completely abolished in vitro editing activity. The carboxy-terminal region of APOBEC1 contains a leucine-rich motif. Deletion analysis of this region indicates that residues 181 to 210 are important for in vitro apoB mRNA editing. Single amino acid substitutions demonstrate that L¹⁸², I¹⁸⁵, and L¹⁸⁹ are important residues required for normal editing function. Furthermore, the double mutant P190A/P191A also lost >90% of editing activity which suggests that a ß turn in this region of the molecule may be essential for proper functioning of APOBEC1. It was suggested that dimerization of APOBEC1 creates an active structure for deamination of apoB mRNA. When we examined the dimerization potential of truncated APOBEC1s using both amino and carboxy termini deletion mutants, we found that amino-terminal deletions up to residue A¹¹⁷ did not impair dimerization activity whereas carboxy-terminal deletions showed diminished dimerization.

The systematic and extensive mutagenesis experiments in this study provide information on the role of various sequence motifs identified in APOBEC1 in enzyme catalysis and dimerization.—Teng, B-B., S. Ochsner, Q. Zhang, K. V. Soman, P. P. Lau, and L. Chan. Mutational analysis of apolipoprotein B mRNA editing enzyme (APOBEC1): structure–function relationships of RNA editing and dimerization. J. Lipid Res. 1999. 40: 623–635.

Supplementary key words: apoB, RNA editing, APOBEC1, cytidine deaminase

CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

This article has been cited by other articles:

				J. Wang, R. Shinkura, M. Muramatsu, H. Nagaoka, K. Kinoshita, and T. Honjo Identification of a Specific Domain Required for Dimerization of Activation-induced Cytidine Deaminase J. Biol. Chem., July 14, 2006; 281(28): 19115 - 19123. [Abstract] [Full Text] [PDF]

				Y.-L. Chiu and W. C. Greene APOBEC3 Cytidine Deaminases: Distinct Antiviral Actions along the Retroviral Life Cycle J. Biol. Chem., March 31, 2006; 281(13): 8309 - 8312. [Abstract] [Full Text] [PDF]

				S. Anant, D. Mukhopadhyay, V. Sankaranand, S. Kennedy, J. O. Henderson, and N. O. Davidson ARCD-1, an apobec-1-related cytidine deaminase, exerts a dominant negative effect on C to U RNA editing Am J Physiol Cell Physiol, December 1, 2001; 281(6): C1904 - C1916. [Abstract] [Full Text] [PDF]

				P. P. Lau, H. Villanueva, K. Kobayashi, M. Nakamuta, B. H.-J. Chang, and L. Chan A DnaJ Protein, Apobec-1-binding Protein-2, Modulates Apolipoprotein B mRNA Editing J. Biol. Chem., November 30, 2001; 276(49): 46445 - 46452. [Abstract] [Full Text] [PDF]

				S. Anant and N. O. Davidson An AU-Rich Sequence Element (UUUN[A/U]U) Downstream of the Edited C in Apolipoprotein B mRNA Is a High-Affinity Binding Site for Apobec-1: Binding of Apobec-1 to This Motif in the 3' Untranslated Region of c-myc Increases mRNA Stability Mol. Cell. Biol., March 15, 2000; 20(6): 1982 - 1992. [Abstract] [Full Text]