Research Update: Molecular and Biochemical Analysis of Phenolic Acid Decarboxylase from Brettanomyces Yeast Isolates

Contamination by Brettanomyces is a major contributor to spoilage of finished wine. One mechanism of spoilage is through metabolism of grape-derived compounds, hydroxycinnamic acids (HCAs). Brettanomyces yeast converts these compounds into volatile products responsible for off-aromas and flavors, notably 4-vinyl phenol (4-VP) and 4-ethyl phenol (4-EP). This “Brett taint” is often described as horse blanket, Band-Aid®, medicinal or earthy. Phenolic acid decarboxylase (PAD) is the first enzyme in the metabolic pathway that converts HCAs to 4-VP and 4-EP. Michael Lentz, associate professor in the Department of Biology at the University of North Florida and his students Jamie Lynn Hall, Rachel Licea and Pricilla Suarez are investigating the role of Brettanomyces metabolism on wine aroma and flavor through molecular analysis of the PAD enzyme.

With funding from an OWB research grant, Lentz and his team have recently acquired three new strains of B. bruxellensis to add to their strain bank. These are isolates from French, Chilean and California wines. They have also added a new B. anomalus strain isolated from Texas Lenoir grapes. These strains have been analyzed for their tolerance to two major HCAs and results indicate that they vary widely. This fits with earlier data that there is significant strain variation in the response to HCAs exposure by Brettanomyces yeast.

The team has also determined the alcohol tolerance of these strains. As generally expected for Brettanomyces, all strains are tolerant to at least 8% ethanol and two of the isolates grew at 20% ethanol, the highest concentration tested.

Lentz and his team have succeeded in cloning the PAD enzyme gene from one of its wine spoilage strains of B. bruxellensis, as well as from a Belgian beer brewing strain. They have initiated experiments to demonstrate that the proteins can be expressed in bacteria. This will allow the team to purify the protein and initiate a more detailed biochemical analysis to better understand the basis for wine spoilage reactions carried out by Brettanomyces yeast.

A better understanding of the spoilage process is essential to reducing the risk associated with these microorganisms and developing new tools to address the economic losses they cause.