Grants and Contributions:

Grant or Award spanning more than one fiscal year. (2017-2018 to 2022-2023)

The Canadian dairy industry heavily relies on the bacterial fermentation for the transformation of milk into products such as cheese and yogurt. An average cheese plant transforms ~1,000,000 litres of milk per day. The starter cultures are a combination of lactic acid bacteria, and Lactococcus lactis is the most important species for cheese manufacture. A cheese plant will rotate different starter cultures, requiring numerous bacterial strains that are carefully selected based on microbiological, biochemical, and technical properties.

In the nonsterile environment of pasteurized milk, the added and preferred bacterial cells will come into contact with ubiquitous virulent phages found in milk. Phages (or bacteriophages) are viruses that infect and replicate within a bacterium. Phage concentration is usually low in milk, but phage population can increase rapidly during the 34 hours cheese making process; virulent lactococcal phages have a 30 min cycle and release of 50-100 virions per infected cell. For decades, the dairy industry has relied on an array of control measures. In spite of these efforts, phages are evolving and variants keep emerging, and phage attacks remain today the most common cause of manufacturing problems in the cheese industry. An in depth understanding of phages will, without a doubt, lead to better strategies for the long-term and optimize control of these phages.

Our primary goal is to significantly increase our knowledge of L. lactis phages that affect the Canadian dairy industry through a characterization of all proteins encoded by these viruses. Even though phages are the most abundant biological entities on the planet (~10 32 phages in the biosphere resulting in ~10 23 infections per second), about 75% of phage genes have unknown functions. Lactococcal phages are classified into ten distinct groups. The 936-type represent 90% of lactococcal phages found in dairy manufacturing facilities and they are the most troublesome. To date, 56 complete genomes of 936-type phages are available. These dsDNA phages encode for few proteins, most of which have unknown functions.

Using a distinct and global approach, we propose to tackle all 3000+ proteins from all 936-type phages with available genome sequences. Structures are essential to understand protein function. In addition, local dynamics in proteins are omnipresent. These movements directly influence events such as ligand binding and catalysis; hence the study of protein dynamics is also essential to understand function. Hence, we also propose in-depth studies of structure and dynamics of highly conserved proteins selected among the 936-type phages. In the first 5 years of this project, we will combine modern prediction tools, solution NMR and molecular dynamics simulations to gather structural and dynamics data on 936-type phage proteins.