Investigation of endoglucanase selectivity on carboxymethyl cellulose by mass spectrometric techniques
- 236 Downloads
The benefits of applying cellulose selective enzymes as analytical tools for chemical structure characterization of cellulose derivatives have been frequently addressed over the years. In a recent study the high selectivity of cellulase Cel45A from Trichoderma reesei (Tr Cel45A) was utilized for relating the chemical structure to the flow properties of carboxymethyl cellulose (CMC). However, in order to take full advantage of the enzymatic hydrolysis the enzyme selectivity on the cellulose substrate must be further investigated. Therefore, the selectivity of Tr Cel45A on CMC was studied by chemical sample preparation of the enzyme products followed by mass spectrometric chemical structure characterization. The results strongly suggest that, in accordance with recent studies, also this highly selective endoglucanase is able to catalyze hydrolysis of glucosidic bonds adjacent to mono-substituted anhydroglucose units (AGUs). Furthermore, the results also indicate that substituents on the nearby AGUs will affect the hydrolysis.
KeywordsCarboxymethyl cellulose Selective hydrolysis Cellulase Permethylation MALDI ESI Mass spectrometry
The authors thank CP Kelco for supplying carboxymethyl cellulose.
- Adden R, Melander C, Brinkmalm G, Gorton L, Mischnick P (2006) New approaches to the analysis of enzymatically hydrolyzed methyl cellulose. Part 1. Investigation of the influence of structural parameters on the extent of degradation. Biomacromolecules 7:1399–1409. doi: 10.1021/bm050941+ CrossRefGoogle Scholar
- Andersson M, Wittgren B, Schagerlof H, Momcilovic D, Wahlund K-G (2004) Size and structure characterization of ethylhydroxyethyl cellulose by the combination of field-flow fractionation with other techniques. Investigation of ultra large components. Biomacromolecules 5:97–105. doi: 10.1021/bm030051z CrossRefGoogle Scholar
- Cohen A, Schagerlof H, Nilsson C, Melander C, Tjerneld F, Gorton L (2004) Liquid chromatography–mass spectrometry analysis of enzyme-hydrolysed carboxymethylcellulose for investigation of enzyme selectivity and substituent pattern. J Chromatogr A 1029:87–95. doi: 10.1016/j.chroma.2003.12.010 CrossRefGoogle Scholar
- Davies GJ, Wilson KS, Henrissat B (1997) Nomenclature for sugar-binding subsites in glycosyl hydrolases. Biochem J 321:557–559Google Scholar
- Fitzpatrick F, Schagerlof H, Andersson T, Richardson S, Tjerneld F, Wahlund K-G et al (2006) NMR, cloud-point measurements and enzymatic depolymerization: complementary tools to investigate substituent patterns in modified celluloses. Biomacromolecules 7:2909–2917. doi: 10.1021/bm060281o CrossRefGoogle Scholar
- Karlsson J, Momcilovic D, Wittgren B, Schülein M, Tjerneld F, Brinkmalm G (2002a) Enzymatic degradation of carboxymethyl cellulose hydrolyzed by the endoglucanases Cel5A, Cel7B, and Cel45A from Humicola insolens and Cel7B, Cel12A and Cel45Acore from Trichoderma reesei. Biopolymers 63:32–40. doi: 10.1002/bip.1060 CrossRefGoogle Scholar
- Momcilovic D, Wittgren B, Wahlund K-G, Karlsson J, Brinkmalm G (2003a) Sample preparation effects in matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry of partially depolymerised carboxymethyl cellulose. Rapid Commun Mass Spectrom 17:1107–1115. doi: 10.1002/rcm.1032 CrossRefGoogle Scholar
- Momcilovic D, Wittgren B, Wahlund K-G, Karlsson J, Brinkmalm G (2003b) Sample preparation effects in matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry of partially depolymerised methyl cellulose. Rapid Commun Mass Spectrom 17:1116–1124. doi: 10.1002/rcm.1033 CrossRefGoogle Scholar
- Richtzenhain H, Lindgren BO, Abrahamsson B, Holmberg K (1954) The alkaline degradation of polysaccharides. I. Degradation of cotton cellulose. Sven Papperstidning 57:363–366Google Scholar
- Saake B, Horner S, Kruse T, Puls J, Liebert T, Heinze T (2000) Detailed investigation on the molecular structure of carboxymethyl cellulose with unusual substitution pattern by means of an enzyme-supported analysis. Macromol Chem Phys 201:1996–2002. doi:10.1002/1521-3935(20001001)201:15<;1996::AID-MACP1996>;3.0.CO;2-XCrossRefGoogle Scholar
- Wirick MG (1968) A study of the enzymic degradation of CMC and other cellulose ethers. J Polym Sci 6:1965–1974Google Scholar