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 |
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Authors: | Sheng Dong Tian-Di Wei Xiu-Lan Chen Chun-Yang Li Peng Wang Bin-Bin Xie Qi-Long Qin Xi-Ying Zhang Xiu-Hua Pang Bai-Cheng Zhou Yu-Zhong Zhang |
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Institution: | From the ‡State Key Laboratory of Microbial Technology and ;the §Marine Biotechnology Research Center, Shandong University, Jinan 250100, China |
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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. Tyr353 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. |
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Keywords: | Alginate Lyase Carbohydrate Metabolism Crystal Structure Docking Mutagenesis Alginate Depolymerization Catalytic Mechanism N-terminal Extension Function Polysaccharide Lyase Family 18 |
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