title: An enhanced protein-protein interaction based on enzymatic complex through replacement of the recognition site.
abstract: Clostridium cellulovorans, produce multi-enzymatic complexes known as cellulosomes, which assemble via the interaction of a dockerin module in the cellulosomal subunit with one of the several cohesin modules in the scaffolding protein, to degrade the plant cell wall polymer. An enhanced cohesin-dockerin interaction was demonstrated by modified certain cellulosomal enzymes with altered amino acid residues at the crucial binding site, 11th and 12th positions in dockerin module. In fluorescence intensity analyses using the cellulosome-based biomarker system, the modified cellulosomal enzymes (EngE SL to AI and EngH SM to AI) showed an increased intensity (1.4- to 2.2-fold) compared with the wild-type proteins. Conversely, modified ExgS (AI to SM) exhibited a reduced intensity (0.6- to 0.7-fold) compared with the wild type. In enzyme-linked and competitive enzyme-linked interaction assays, the some modified protein (EngE SL to AI and EngH SM to AI) showed their increased binding affinity toward the cohesins (Coh2 and Coh9). Surface plasmon resonance analysis quantitatively demonstrated the binding affinity of these two modified proteins toward cohesins showed similar or higher affinity comparing with its with wild type proteins. These results suggest the replacement of amino acid residues in the certain recognition site significantly affects the binding affinity of the cohesin-dockerin interaction.
authors.affiliation: Department of Biotechnology, Korea University, Seoul 136-701, Republic of Korea.
authors.forename: Sang Duck
authors.initials: SD
authors.lastname: Jeon
authors.affiliation: Department of Biotechnology, Korea University, Seoul 136-701, Republic of Korea.
authors.forename: Su Jung
authors.initials: SJ
authors.lastname: Kim
authors.affiliation: Department of Biotechnology, Korea University, Seoul 136-701, Republic of Korea.
authors.forename: Sung Hyun
authors.initials: SH
authors.lastname: Park
authors.affiliation: Changhae Advanced Institute of Technology, Changhae Ethanol Co., Ltd., Jeonju 561-203, Republic of Korea.
authors.forename: Gi-Wook
authors.initials: GW
authors.lastname: Choi
authors.affiliation: Department of Biotechnology, Korea University, Seoul 136-701, Republic of Korea. Electronic address: samhan@korea.ac.kr.
authors.forename: Sung Ok
authors.initials: SO
authors.lastname: Han
mesh_terms: D015415:Biomarkers / Q000378:metabolism; D018797:Cell Cycle Proteins; D044603:Cellulosomes / Q000378:metabolism; D002868:Chromosomal Proteins, Non-Histone; D003013:Clostridium / Q000201:enzymology*; D015180:Electrophoresis, Gel, Two-Dimensional; D005453:Fluorescence; D007700:Kinetics; D009097:Multienzyme Complexes / Q000378:metabolism*; D011485:Protein Binding; D020349:Surface Plasmon Resonance; D000097722:Cohesins
chemical_list: D015415:Biomarkers; D018797:Cell Cycle Proteins; D002868:Chromosomal Proteins, Non-Histone; D009097:Multienzyme Complexes; D000097722:Cohesins
keywords: Cellulosome; Clostridium cellulovorans; Cohesin–dockerin interaction