K. SAEKI (1), A. Suyama (2), K. Takegawa (2), M. Sato (1), T. Shigyo (1); (1) Sapporo Breweries Ltd., Shizuoka, Japan; (2) Kyushu University, Fukuoka, Japan

Analytical
Friday, June 6 - 2:00 p.m.-3:45 p.m.
Level 4, Red Lacquer Ballroom

Foam is one of the most important properties of beer. Many factors, including barley and hop varieties, malting conditions, and yeast strain, influence foam production and stability. There are also factors that negatively affect foam quality, such as lipids, alcohol, and proteases produced by yeast. Yeast-derived vacuolar protease proteinase A (PrA) digests proteins in beer and, thus, impairs foam stability. “Vacuolar” protein PrA is normally transferred to vacuoles from the Golgi, but under specific conditions during fermentation, PrA leaves the cell for some reason, which is detrimental to the final product. In the brewing industry, PrA has been of great interest for more than 40 years. Methods of measuring PrA activity in beer have been established, and several trials have been performed to reduce the amount of PrA excreted in beer. However, the fundamental mechanism causing PrA to leave the cell has yet to be elucidated. Therefore, we conducted assays to monitor the behavior of PrA and its related proteins both inside and outside of the cell, aiming to identify their relationships in the intracellular transport system. PrA is mainly transported to vacuoles via Vps10p, a vacuolar protein receptor located in late Golgi. Vps10p is also involved in the transport of another vacuolar protease, carboxypeptidase Y (CPY), so we hypothesized that the change in the ratio of these three proteins would be crucial for the irregular sorting of PrA toward the outside of the cell. To demonstrate this, we performed western analysis and protein activity assay to analyze the amount of each protein both inside and outside the cell. Fermentation tests were also conducted using various kinds of wort with different compositions to reveal its effect on PrA activity in beer. Analysis with yeast in shaking culture indicated that different growth conditions cause changes in the amount and activity of each protein and that they also affect the ratio of these three proteins. Based on these results, we proposed a model to describe how PrA leaves the cell.

Kei Saeki received a master’s degree from the Department of Biotechnology, University of Tokyo, Japan. She began her career as a microbiologist in 2012 at the Frontier Laboratories of Value Creation, Sapporo Breweries Ltd. In 2013, she started her studies in intracellular transport supervised by Professor Takegawa at the Department of Bioscience and Biotechnology, Kyushu University, Japan.