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. 2022 Jun 1;12:9135. doi: 10.1038/s41598-022-13124-1

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© The Author(s) 2022

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

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Experiment-3 of the intestinal pancreatic trypsin (78 U/mL) effects (with describing model parameter estimates $\pm SE)$ on heat-treated GH5-tCel5A1 activity over a 5-h period after the intestinal pancreatic trypsin buffer being mixed with the heat-treated GH5-tCel5A1 enzyme preparation for incubation at the resulting mixture pH = 6.0–7.0 and trypsin (78 U/mL) prior to further incubation for measuring cellulase activity with the carboxymethyl cellulose (CMC) substrate buffer in presence of dithiothreitol (DTT) (5 mM) and N₂ gas purging for 15 min at 37 °C. (A) Plot of the inhibition kinetic relationship between the heat-treated GH5-tCel5A1 activity (v, mean $\pm SE, n = 12 at each time point,$ nmol/mg protein•min) and incubation time (t, h) under the trypsin (78 U/mL) condition established according to the quadratic response model as: $Y = 17.408 (\pm 1.388) - 8.694 (\pm 1.154) x + 1.259 (\pm 0.198) x^{2}$ , P < 0.0001 for all parameter estimates; $R^{2} = 0.475$ , n = 84; and alternatively according to the linear response model established as $y = 10.718 (\pm 1.103) - 1.599 (\pm 0.357) x$ , P < 0.0001 for all parameter estimates, $r^{2} = 0.209$ , n = 84; (B) Quadratic plot of the inhibition kinetic relationship between the relationship of relative heat-treated GH5-tCel5A1 activity (v, mean $\pm SE, n = 12 at each time point,$ % of the control group measured at I_c of 100.000 ± 9.682) at 0-min exposure to the trypsin-78-U/mL incubation) and incubation time trypsin (78 U/mL) conditions established according to the quadratic response model as: $Y = 114.3 (\pm 9.116) - 57.093 (\pm 7.581) x + 8.273 (\pm 1.302) x^{2}$ , P < 0.0001 for all parameter estimates, $R^{2} = 0.475$ , n = 84; and alternatively established according to the linear response model as $y = 70.388 (\pm 7.242) - 10.502 (\pm 2.346) x$ , P < 0.0001 for all parameter estimates, $r^{2} = 0.210$ , n = 84; (C) the Eadie-Hofstee linear plot between the heat-treated GH5-tCel5A1 enzyme activity (v, mean $\pm SE, n = 12 at each data point,$ nmol/mg protein•min) and the v/t ratio established as $y = 1.918 (\pm 0.353) + 1.322 (\pm 0.092) x$ , P < 0.0001 for all parameter estimates; $r^{2} = 0.749$ , n = 72; and (D) the Eadie-Hofstee linear plot between the relative heat-treated GH5-tCel5A1 enzyme activity (v, mean $\pm SE, n = 12 at each data point,$ % of the control group measured at I_c of 100.000 ± 9.682) at 0-min exposure to the trypsin-78-U/mL incubation) and the v/t ratio established as $y = 12.602 (\pm 2.321) + 1.322 (\pm 0.092) x$ , P < 0.0001 for all parameter estimates; $r^{2} = 0.749$ , n = 72.