The fumR gene encoding fumarase in the filamentous fungus Rhizopus oryzae: cloning, structure and expression

Area of Science:

• Biochemistry
• Molecular Biology
• Mycology

Background:

• Filamentous fungus Rhizopus oryzae (Ro) is known for high fumaric acid (FA) accumulation.
• Understanding the molecular basis of FA overproduction is crucial.

Purpose of the Study:

• To investigate the molecular mechanisms behind increased fumaric acid biosynthesis in R. oryzae.
• To clone and analyze the structure and expression of the fumarase gene (fumR).

Main Methods:

• Gene cloning and nucleotide sequencing of fumR.
• Bioinformatic analysis of the fumR gene and its product.
• Protein detection using immunoassays.
• RNA transcript analysis via primer extension and Northern blotting.

Main Results:

• The fumR gene contains nine introns, encoding a 494-amino acid class-II fumarase.
• A 50 kDa fumarase protein was detected in crude extracts.
• fumR RNA levels significantly increased under stress conditions (high carbon, low nitrogen).
• A single transcript for fumR was identified.

Conclusions:

• Transcriptional regulation of the fumR gene is likely involved in fumaric acid overproduction in R. oryzae under stress.
• Further investigation into additional regulatory mechanisms is warranted.