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Recombinant Expression and Characterization of Two Isoforms of Anopheles gambiae Laccase‐2
Author(s) -
Sullivan Lucinda Ilene,
Gorman Maureen J,
Dittmer Neal T,
Kanost Michael R
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.21.6.a999
Laccases are multicopper oxidases that act on a broad range of substrates. Laccases in fungi, plants, and bacteria have roles in detoxification, pigmentation, wound healing, and morphogenesis. Insects have multiple laccase genes, but the function of just one type is known; laccase‐2 orthologs are required for tanning of newly synthesized exoskeleton. We have been studying the role of Lac2 in a mosquito species, Anopheles gambiae . In all insect species whose genomes have been sequenced, alternative exon splicing generates two isoforms of Lac2. The objective of this study was to characterize the two isoforms of AgLac2. They are identical in their first 500 residues, but the carboxyl‐terminal 262 residues derived from alternative exons are 81% identical. The A and B isoforms are expressed in all developmental stages, but their relative abundance differs, and only the B isoform is upregulated in ovaries following a blood meal. Recombinant Lac2A and Lac2B were expressed and purified. They are both glycoproteins of ~80 kDa, and both can oxidize the laccase substrate ABTS. Preliminary results indicate that Lac2A and Lac2B have pH optima of 4.0–4.5 and 3.5, respectively. Although the two isoforms are very similar in their amino acid sequences, differences in their expression and catalytic properties may indicate different roles in insect physiology. Supported by NIH AI057815.