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J Insect Sci. 2007;7:1-13. doi: 10.1673/031.007.6201.

Porphobilinogen synthase from the butterfly, Pieris brassicae: purification and comparative characterization.

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1
Department of General Zoology and Endocrinology (Biology I), University of Ulm, Albert-Einstein-Allee 11, D-89069 Ulm, Germany.

Abstract

Porphobilinogen represents a key building block of tetrapyrroles serving as functional ligands of many vitally important proteins. Here we report the first purification of porphobilinogen synthase (PBGS) from whole insects by sequentially employing two modes of native electrophoresis on polyacrylamide gels subsequent to more conventional procedures. Using adults of Pieris brassicae L. (Lepidoptera: Pieridae) we achieved approximately 10,000-fold purification with final yields of up to 25% of electrophoretically pure PBGS with a specific activity of approximately 160 micromol PBG h(-1) mg(-1) at 37 degrees C and an affinity of 0.36 mM to its substrate 5-aminolevulinic acid. Enzyme activity was inhibited by the substrate mimics, levulinic acid and succinylacetone, and by chelating agents. PBGS behaved as a relatively heat-stable octameric complex of 292.3 kDa composed of 36.5 kDa subunits. Most general features of this insect PBGS were comparable to those published for other animal PBGS enzymes, while remarkable differences were found to the reported recombinant Drosophila enzyme. Moreover, rabbit antiserum directed against purified Pieris PBGS revealed significant immunological differences among insect PBGS enzymes from a wide range of orders contrasting to the overall evolutionary conserved features of this enzyme.

PMID:
20302542
PMCID:
PMC2999460
DOI:
10.1673/031.007.6201
[Indexed for MEDLINE]
Free PMC Article
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