G T Howard1, J Vicknair, R I MacKie. 1. Department of Biological Sciences, South-eastern Louisiana University, Hammond, LA 70402, USA.
Abstract
AIMS: A plate assay to screen and detect bacterial polyurethanase in agar medium containing a colloidal polyester-polyurethane and rhodamine B is presented. METHODS AND RESULTS: Substrate hydrolysis causes the formation of orange fluorescent halos visible upon u.v. irradiation. The logarithm of polyurethanase activity from a purified polyurethanase protein is linearly correlated with the diameter of halos, thereby allowing quantification of polyurethanase activities ranging from 0.81 to 7.29 Units. CONCLUSIONS: The potential advantages of this system are in identification and recovery of viable polyurethanolytic bacteria and quantification of polyurethanase activity. SIGNIFICANCE AND IMPACT OF THE STUDY: These advantages are derived largely from the intense fluorescence observed due to the hydrolysis of substrate reacting with rhodamine B allowing for the use of low substrate concentrations and corresponding decrease in time required detecting low levels of enzyme activity.
AIMS: A plate assay to screen and detect bacterial polyurethanase in agar medium containing a colloidal polyester-polyurethane and rhodamine B is presented. METHODS AND RESULTS: Substrate hydrolysis causes the formation of orange fluorescent halos visible upon u.v. irradiation. The logarithm of polyurethanase activity from a purified polyurethanase protein is linearly correlated with the diameter of halos, thereby allowing quantification of polyurethanase activities ranging from 0.81 to 7.29 Units. CONCLUSIONS: The potential advantages of this system are in identification and recovery of viable polyurethanolytic bacteria and quantification of polyurethanase activity. SIGNIFICANCE AND IMPACT OF THE STUDY: These advantages are derived largely from the intense fluorescence observed due to the hydrolysis of substrate reacting with rhodamine B allowing for the use of low substrate concentrations and corresponding decrease in time required detecting low levels of enzyme activity.
Authors: Jonathan R Russell; Jeffrey Huang; Pria Anand; Kaury Kucera; Amanda G Sandoval; Kathleen W Dantzler; DaShawn Hickman; Justin Jee; Farrah M Kimovec; David Koppstein; Daniel H Marks; Paul A Mittermiller; Salvador Joel Núñez; Marina Santiago; Maria A Townes; Michael Vishnevetsky; Neely E Williams; Mario Percy Núñez Vargas; Lori-Ann Boulanger; Carol Bascom-Slack; Scott A Strobel Journal: Appl Environ Microbiol Date: 2011-07-15 Impact factor: 4.792