Assay Replicability in β-Glucosidase Enzyme Kinetics Across Laboratories

Abstract

Replicability is the foundation of research in any scientific discipline. Despite this fact, few studies address experimental variability within and across multiple institutions that operate under the same protocol. While consistency is usually well documented within the same lab, multi-institutional experiments may introduce new variables and, therefore, variability that may lead to inconsistent results. This study seeks to explore intra- and interinstitutional variability among enzyme catalytic efficiency values (KM and Kcat/KM) for the wild type of β-Glucosidase derived from Paenibacillus polymyxa. A standardized protocol for the assay was provided to all institutions that participated in the study. The analysis was conducted using data from 13 laboratories across the United States. The information was collected through the Design2Data CURE database. A total of 132 independent assays of β-Glucosidase were analyzed. Statistical analysis of Kcat/KM , KM, and T50 was conducted using SPSS, and Whisker Plots with a 90% confidence interval were generated for all parameters. Acceptance intervals for the parameters assayed of β-Glucosidase were determined; these could be used in all future experiments in the network as a reference by current and incoming laboratory members that incorporate into the network yearly. Ultimately, we aim to identify possible errors and misunderstandings in the protocol we use in the laboratories of the D2D network to improve assay replicability across institutions. High variability may be a concern when data is interpreted across institutions. This specific assay is used as a control for other, more complex assays in the CURE network and, therefore, the variability found can potentially lead to incorrect interpretation of results. It also highlights the high variability of a simple enzymatic assay when it is replicated in laboratories with different personnel and equipment, regardless of using the same standardized protocol.