title: Molecular insight into the role of the N-terminal extension in the maturation, substrate recognition, and catalysis of a bacterial alginate lyase from polysaccharide lyase family 18.
abstract: Bacterial alginate lyases, which are members of several polysaccharide lyase (PL) families, have important biological roles and biotechnological applications. The mechanisms for maturation, substrate recognition, and catalysis of PL18 alginate lyases are still largely unknown. A PL18 alginate lyase, aly-SJ02, from Pseudoalteromonas sp. 0524 displays a β-jelly roll scaffold. Structural and biochemical analyses indicated that the N-terminal extension in the aly-SJ02 precursor may act as an intramolecular chaperone to mediate the correct folding of the catalytic domain. Molecular dynamics simulations and mutational assays suggested that the lid loops over the aly-SJ02 active center serve as a gate for substrate entry. Molecular docking and site-directed mutations revealed that certain conserved residues at the active center, especially those at subsites +1 and +2, are crucial for substrate recognition. Tyr(353) may function as both a catalytic base and acid. Based on our results, a model for the catalysis of aly-SJ02 in alginate depolymerization is proposed. Moreover, although bacterial alginate lyases from families PL5, 7, 15, and 18 adopt distinct scaffolds, they share the same conformation of catalytic residues, reflecting their convergent evolution. Our results provide the foremost insight into the mechanisms of maturation, substrate recognition, and catalysis of a PL18 alginate lyase.
authors.affiliation: From the State Key Laboratory of Microbial Technology and the Marine Biotechnology Research Center, Shandong University, Jinan 250100, China.
authors.forename: Sheng
authors.initials: S
authors.lastname: Dong
authors.affiliation: From the State Key Laboratory of Microbial Technology and.
authors.forename: Tian-Di
authors.initials: TD
authors.lastname: Wei
authors.affiliation: From the State Key Laboratory of Microbial Technology and the Marine Biotechnology Research Center, Shandong University, Jinan 250100, China.
authors.forename: Xiu-Lan
authors.initials: XL
authors.lastname: Chen
authors.affiliation: From the State Key Laboratory of Microbial Technology and the Marine Biotechnology Research Center, Shandong University, Jinan 250100, China.
authors.forename: Chun-Yang
authors.initials: CY
authors.lastname: Li
authors.affiliation: From the State Key Laboratory of Microbial Technology and the Marine Biotechnology Research Center, Shandong University, Jinan 250100, China.
authors.forename: Peng
authors.initials: P
authors.lastname: Wang
authors.affiliation: From the State Key Laboratory of Microbial Technology and the Marine Biotechnology Research Center, Shandong University, Jinan 250100, China.
authors.forename: Bin-Bin
authors.initials: BB
authors.lastname: Xie
authors.affiliation: From the State Key Laboratory of Microbial Technology and the Marine Biotechnology Research Center, Shandong University, Jinan 250100, China.
authors.forename: Qi-Long
authors.initials: QL
authors.lastname: Qin
authors.affiliation: From the State Key Laboratory of Microbial Technology and the Marine Biotechnology Research Center, Shandong University, Jinan 250100, China.
authors.forename: Xi-Ying
authors.initials: XY
authors.lastname: Zhang
authors.affiliation: From the State Key Laboratory of Microbial Technology and the Marine Biotechnology Research Center, Shandong University, Jinan 250100, China.
authors.forename: Xiu-Hua
authors.initials: XH
authors.lastname: Pang
authors.affiliation: From the State Key Laboratory of Microbial Technology and the Marine Biotechnology Research Center, Shandong University, Jinan 250100, China.
authors.forename: Bai-Cheng
authors.initials: BC
authors.lastname: Zhou
authors.affiliation: From the State Key Laboratory of Microbial Technology and the Marine Biotechnology Research Center, Shandong University, Jinan 250100, China zhangyz@sdu.edu.cn.
authors.forename: Yu-Zhong
authors.initials: YZ
authors.lastname: Zhang
mesh_terms: D000595:Amino Acid Sequence; D000596:Amino Acids / Q000378:metabolism; D055162:Biocatalysis; D020134:Catalytic Domain; D002942:Circular Dichroism; D003198:Computer Simulation; D018360:Crystallography, X-Ray; D008958:Models, Molecular; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism; D011133:Polysaccharide-Lyases / Q000737:chemistry* / Q000378:metabolism*; D017433:Protein Structure, Secondary; D031161:Pseudoalteromonas / Q000201:enzymology*; D020539:Sequence Analysis, Protein; D040681:Structural Homology, Protein; D013329:Structure-Activity Relationship; D013379:Substrate Specificity
chemical_list: D000596:Amino Acids; D050505:Mutant Proteins; D011133:Polysaccharide-Lyases; C016158:poly(beta-D-mannuronate) lyase
keywords: Alginate Depolymerization; Alginate Lyase; Carbohydrate Metabolism; Catalytic Mechanism; Crystal Structure; Docking; Mutagenesis; N-terminal Extension Function; Polysaccharide Lyase Family 18