Liquid chromatography combined with mass spectrometry for the investigation of endoglucanase selectivity on carboxymethyl cellulose.
Endoglucanases are useful tools in the chemical structure analysis of cellulose derivatives. However, knowledge on the endoglucanase selectivity, which is of central importance for data interpretation, is still limited. In this study, new reverse-phase liquid chromatography mass spectrometry (LC-MS) methods were developed to investigate the selectivity of the endoglucanases Cel5A, Cel7B, Cel45A, and Cel74A from the filamentous fungus Trichoderma reesei. The aim was to improve the identification of the regioisomers in the complex mixtures that are obtained after enzymatic hydrolysis. Reduction followed by per-O-methylation was performed in order to improve the separation in reverse-phase LC, increase MS sensitivity, and to facilitate structure analysis by MS/MS of O-carboxymethyl glucose and cellooligosaccharides. The cellulose selective enzymes that were investigated displayed interesting differences in enzyme selectivity on CMC substrates.
Enebro
Jonas
J
School of Chemical Science and Engineering, Fibre and Polymer Technology, Royal Institute of Technology (KTH), Stockholm, Sweden.
Momcilovic
Dane
D
Siika-Aho
Matti
M
Karlsson
Sigbritt
S
D002266:Carboxymethylcellulose Sodium / Q000737:chemistry / Q000378:metabolism*; D002480:Cellulase / Q000378:metabolism*; D002853:Chromatography, Liquid; D045424:Complex Mixtures / Q000032:analysis / Q000737:chemistry / Q000378:metabolism; D006868:Hydrolysis; D013058:Mass Spectrometry; D013237:Stereoisomerism; D013379:Substrate Specificity; D053719:Tandem Mass Spectrometry; D014242:Trichoderma / Q000201:enzymology
D045424:Complex Mixtures; D002480:Cellulase; D002266:Carboxymethylcellulose Sodium