Intracellular Beta-carbonic Anhydrase of the Unicellular Green Alga Coccomyxa. Cloning of the Cdna and Characterization of the Functional Enzyme Overexpressed in Escherichia Coli

Overview

Journal Plant Physiol

Specialty Physiology

Date 1998 Aug 14

PMID 9701590

Citations 10

Authors

T Hiltonen

H Bjorkbacka

C Forsman

A K Clarke

G Samuelsson

Affiliations

Soon will be listed here.

Abstract

Carbonic anhydrase (CA) (EC 4.2.1.1) enzymes catalyze the reversible hydration of CO2, a reaction that is important in many physiological processes. We have cloned and sequenced a full-length cDNA encoding an intracellular beta-CA from the unicellular green alga Coccomyxa. Nucleotide sequence data show that the isolated cDNA contains an open reading frame encoding a polypeptide of 227 amino acids. The predicted polypeptide is similar to beta-type CAs from Escherichia coli and higher plants, with an identity of 26% to 30%. The Coccomyxa cDNA was overexpressed in E. coli, and the enzyme was purified and biochemically characterized. The mature protein is a homotetramer with an estimated molecular mass of 100 kD. The CO2-hydration activity of the Coccomyxa enzyme is comparable with that of the pea homolog. However, the activity of Coccomyxa CA is largely insensitive to oxidative conditions, in contrast to similar enzymes from most higher plants. Fractionation studies further showed that Coccomyxa CA is extrachloroplastic.

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