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Appl. Environ. Microbiol., Sep 1997, 3488-3493, Vol 63, No. 9
SJ Rasmussen-Wilson, JS Palas, VJ Wolf, CS Taft and CP Selitrennikoff
Heterologous expression of plant genes may serve as an important
alternative for producing plant proteins. We have investigated the ability
of the fungus Neurospora crassa to secrete zeamatin, a protein produced by
Zea mays. Zeamatin was induced after being fused to glucoamylase, an
extracellular hydrolase produced by N. crassa. Glucoamylase induction and
other culture parameters were monitored in untransformed N. crassa grown in
shaken liquid culture. A DNA plasmid, pGEZ, was constructed by inserting
zeamatin-encoding cDNA into an expression cassette containing the promoter,
a truncated open reading frame, and the terminator sequence of the N.
crassa glucoamylase gene. Zeamatin-encoding cDNA was modified at the N
terminus to include a kex- 2 protease site, allowing cleavage of the
chimeric product in the secretory pathway. Strains containing the chimeric
gene construct were grown in liquid culture and induced for glucoamylase
and zeamatin production. Zeamatin antibody detected a protein in a Western
blot of concentrated culture supernatants that comigrated with authentic
zeamatin. Secreted zeamatin was active in inhibiting the growth of Candida
albicans in an agar diffusion assay, indicating that zeamatin had been
correctly synthesized, processed, and secreted by N. crassa.
Copyright © 1997, American Society for Microbiology
Expression of a plant protein by Neurospora crassa
MycoTox, Inc., Denver, Colorado 80262, USA. swilso@mycotox.abti.com
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