Adnan Kadić1, Anikó Várnai2, Vincent G H Eijsink2, Svein Jarle Horn3, Gunnar Lidén4. 1. Department of Chemical Engineering, Lund University, P.O. Box 118, 221 00, Lund, Sweden. 2. Faculty of Chemistry, Biotechnology and Food Sciences, Norwegian University of Life Sciences (NMBU), P.O. Box 5003, NO-1432, Ås, Norway. 3. Faculty of Chemistry, Biotechnology and Food Sciences, Norwegian University of Life Sciences (NMBU), P.O. Box 5003, NO-1432, Ås, Norway. svein.horn@nmbu.no. 4. Department of Chemical Engineering, Lund University, P.O. Box 118, 221 00, Lund, Sweden. gunnar.liden@chemeng.lth.se.
Abstract
BACKGROUND: Biochemical conversion of lignocellulosic biomass to simple sugars at commercial scale is hampered by the high cost of saccharifying enzymes. Lytic polysaccharide monooxygenases (LPMOs) may hold the key to overcome economic barriers. Recent studies have shown that controlled activation of LPMOs by a continuous H2O2 supply can boost saccharification yields, while overdosing H2O2 may lead to enzyme inactivation and reduce overall sugar yields. While following LPMO action by ex situ analysis of LPMO products confirms enzyme inactivation, currently no preventive measures are available to intervene before complete inactivation. RESULTS: Here, we carried out enzymatic saccharification of the model cellulose Avicel with an LPMO-containing enzyme preparation (Cellic CTec3) and H2O2 feed at 1 L bioreactor scale and followed the oxidation-reduction potential and H2O2 concentration in situ with corresponding electrode probes. The rate of oxidation of the reductant as well as the estimation of the amount of H2O2 consumed by LPMOs indicate that, in addition to oxidative depolymerization of cellulose, LPMOs consume H2O2 in a futile non-catalytic cycle, and that inactivation of LPMOs happens gradually and starts long before the accumulation of LPMO-generated oxidative products comes to a halt. CONCLUSION: Our results indicate that, in this model system, the collapse of the LPMO-catalyzed reaction may be predicted by the rate of oxidation of the reductant, the accumulation of H2O2 in the reactor or, indirectly, by a clear increase in the oxidation-reduction potential. Being able to monitor the state of the LPMO activity in situ may help maximizing the benefit of LPMO action during saccharification. Overcoming enzyme inactivation could allow improving overall saccharification yields beyond the state of the art while lowering LPMO and, potentially, cellulase loads, both of which would have beneficial consequences on process economics.
BACKGROUND: Biochemical conversion of lignocellulosic biomass to simple sugars at commercial scale is hampered by the high cost of saccharifying enzymes. Lytic polysaccharide monooxygenases (LPMOs) may hold the key to overcome economic barriers. Recent studies have shown that controlled activation of LPMOs by a continuous H2O2 supply can boost saccharification yields, while overdosing H2O2 may lead to enzyme inactivation and reduce overall sugar yields. While following LPMO action by ex situ analysis of LPMO products confirms enzyme inactivation, currently no preventive measures are available to intervene before complete inactivation. RESULTS: Here, we carried out enzymatic saccharification of the model cellulose Avicel with an LPMO-containing enzyme preparation (Cellic CTec3) and H2O2 feed at 1 L bioreactor scale and followed the oxidation-reduction potential and H2O2 concentration in situ with corresponding electrode probes. The rate of oxidation of the reductant as well as the estimation of the amount of H2O2 consumed by LPMOs indicate that, in addition to oxidative depolymerization of cellulose, LPMOs consume H2O2 in a futile non-catalytic cycle, and that inactivation of LPMOs happens gradually and starts long before the accumulation of LPMO-generated oxidative products comes to a halt. CONCLUSION: Our results indicate that, in this model system, the collapse of the LPMO-catalyzed reaction may be predicted by the rate of oxidation of the reductant, the accumulation of H2O2 in the reactor or, indirectly, by a clear increase in the oxidation-reduction potential. Being able to monitor the state of the LPMO activity in situ may help maximizing the benefit of LPMO action during saccharification. Overcoming enzyme inactivation could allow improving overall saccharification yields beyond the state of the art while lowering LPMO and, potentially, cellulase loads, both of which would have beneficial consequences on process economics.
Authors: Silja Kuusk; Bastien Bissaro; Piret Kuusk; Zarah Forsberg; Vincent G H Eijsink; Morten Sørlie; Priit Väljamäe Journal: J Biol Chem Date: 2017-11-14 Impact factor: 5.157
Authors: Matthias Frommhagen; Martijn J Koetsier; Adrie H Westphal; Jaap Visser; Sandra W A Hinz; Jean-Paul Vincken; Willem J H van Berkel; Mirjam A Kabel; Harry Gruppen Journal: Biotechnol Biofuels Date: 2016-08-31 Impact factor: 6.040
Authors: Brian R Scott; Hong Zhi Huang; Jesper Frickman; Rune Halvorsen; Katja S Johansen Journal: Biotechnol Lett Date: 2015-11-05 Impact factor: 2.461
Authors: Heidi Østby; Line Degn Hansen; Svein J Horn; Vincent G H Eijsink; Anikó Várnai Journal: J Ind Microbiol Biotechnol Date: 2020-08-25 Impact factor: 3.346
Authors: Tobias M Hedison; Erik Breslmayr; Muralidharan Shanmugam; Kwankao Karnpakdee; Derren J Heyes; Anthony P Green; Roland Ludwig; Nigel S Scrutton; Daniel Kracher Journal: FEBS J Date: 2021-01-26 Impact factor: 5.622
Authors: James Li; Ethan D Goddard-Borger; Olanrewaju Raji; Hirak Saxena; Laleh Solhi; Yann Mathieu; Emma R Master; Warren W Wakarchuk; Harry Brumer Journal: Appl Environ Microbiol Date: 2022-07-12 Impact factor: 5.005