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Applied and Environmental Microbiology, August 2004, p. 4814-4820, Vol. 70, No. 8
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.8.4814-4820.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Overexpression of Lactobacillus casei D-Hydroxyisocaproic Acid Dehydrogenase in Cheddar Cheese{dagger}

Jeffery R. Broadbent,1* Sanjay Gummalla,1,{ddagger} Joanne E. Hughes,2 Mark E. Johnson,3 Scott A. Rankin,4 and Mary Anne Drake5

Western Dairy Center, Department of Nutrition and Food Sciences,1 Department of Biology, Utah State University, Logan, Utah 84322,2 Wisconsin Center for Dairy Research,3 Department of Food Science, University of Wisconsin, Madison, Wisconsin 53706,4 Southeast Dairy Foods Research Center and Department of Food Science, North Carolina State University, Raleigh, North Carolina 276955

Received 13 January 2004/ Accepted 5 May 2004

Metabolism of aromatic amino acids by lactic acid bacteria is an important source of off-flavor compounds in Cheddar cheese. Previous work has shown that {alpha}-keto acids produced from Trp, Tyr, and Phe by aminotransferase enzymes are chemically labile and may degrade spontaneously into a variety of off-flavor compounds. However, dairy lactobacilli can convert unstable {alpha}-keto acids to more-stable {alpha}-hydroxy acids via the action of {alpha}-keto acid dehydrogenases such as D-hydroxyisocaproic acid dehydrogenase. To further characterize the role of this enzyme in cheese flavor, the Lactobacillus casei D-hydroxyisocaproic acid dehydrogenase gene was cloned into the high-copy-number vector pTRKH2 and transformed into L. casei ATCC 334. Enzyme assays confirmed that {alpha}-keto acid dehydrogenase activity was significantly higher in pTRKH2:dhic transformants than in wild-type cells. Reduced-fat Cheddar cheeses were made with Lactococcus lactis starter only, starter plus L. casei ATCC 334, and starter plus L. casei ATCC 334 transformed with pTRKH2:dhic. After 3 months of aging, the cheese chemistry and flavor attributes were evaluated instrumentally by gas chromatography-mass spectrometry and by descriptive sensory analysis. The culture system used significantly affected the concentrations of various ketones, aldehydes, alcohols, and esters and one sulfur compound in cheese. Results further indicated that enhanced expression of D-hydroxyisocaproic acid dehydrogenase suppressed spontaneous degradation of {alpha}-keto acids, but sensory work indicated that this effect retarded cheese flavor development.

* Corresponding author. Mailing address: Western Dairy Center, Department of Nutrition and Food Sciences, Utah State University, Logan, UT 84322-8700. Phone: (435) 797-2113. Fax: (435) 797-2379. E-mail: broadbnt{at}cc.usu.edu.

{dagger} Approved as Utah Agricultural Experiment Station journal paper number 7488.

{ddagger} Present address: Degussa Flavors and Fruit Systems US, LLC, Cincinnati, OH 45215.

Applied and Environmental Microbiology, August 2004, p. 4814-4820, Vol. 70, No. 8
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.8.4814-4820.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



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