title: Surface Immobilization of Human Arginase-1 with an Engineered Ice Nucleation Protein Display System in E. coli.
abstract: Ice nucleation protein (INP) is frequently used as a surface anchor for protein display in gram-negative bacteria. Here, MalE and TorA signal peptides, and three charged polypeptides, 6×Lys, 6×Glu and 6×Asp, were anchored to the N-terminus of truncated INP (InaK-N) to improve its surface display efficiency for human Arginase1 (ARG1). Our results indicated that the TorA signal peptide increased the surface translocation of non-protein fused InaK-N and human ARG1 fused InaK-N (InaK-N/ARG1) by 80.7% and 122.4%, respectively. Comparably, the MalE signal peptide decreased the display efficiencies of both the non-protein fused InaK-N and InaK-N/ARG1. Our results also suggested that the 6×Lys polypeptide significantly increased the surface display efficiency of K6-InaK-N/ARG1 by almost 2-fold, while also practically abolishing the surface translocation of non-protein fused InaK-N, indicating the interesting roles of charged polypeptides in bacteria surface display systems. Cell surface-immobilized K6-InaK-N/ARG1 presented an arginase activity of 10.7 U/OD600 under the optimized conditions of 40°C, pH 10.0 and 1 mM Mn2+, which could convert more than 95% of L-Arginine (L-Arg) to L-Ornithine (L-Orn) in 16 hours. The engineered InaK-Ns expanded the INP surface display system, which aided in the surface immobilization of human ARG1 in E. coli cells.
authors.affiliation: Hubei Collaborative Innovation Center for Green Transformation of Bio-resources, Hubei Key Laboratory of Industrial Biotechnology, College of Life Sciences, Hubei University, Wuhan, People's Republic of China.
authors.forename: Zhen
authors.initials: Z
authors.lastname: Zhang
authors.affiliation: Hubei Collaborative Innovation Center for Green Transformation of Bio-resources, Hubei Key Laboratory of Industrial Biotechnology, College of Life Sciences, Hubei University, Wuhan, People's Republic of China.
authors.forename: Rongxin
authors.initials: R
authors.lastname: Tang
authors.affiliation: Beihai Blood Center, Guang Xi, People's Republic of China.
authors.forename: Lu
authors.initials: L
authors.lastname: Bian
authors.affiliation: Hubei Collaborative Innovation Center for Green Transformation of Bio-resources, Hubei Key Laboratory of Industrial Biotechnology, College of Life Sciences, Hubei University, Wuhan, People's Republic of China.
authors.forename: Meng
authors.initials: M
authors.lastname: Mei
authors.affiliation: Hubei Collaborative Innovation Center for Green Transformation of Bio-resources, Hubei Key Laboratory of Industrial Biotechnology, College of Life Sciences, Hubei University, Wuhan, People's Republic of China.
authors.forename: Chunhua
authors.initials: C
authors.lastname: Li
authors.affiliation: Hubei Collaborative Innovation Center for Green Transformation of Bio-resources, Hubei Key Laboratory of Industrial Biotechnology, College of Life Sciences, Hubei University, Wuhan, People's Republic of China.
authors.forename: Xiangdong
authors.initials: X
authors.lastname: Ma
authors.affiliation: Hubei Collaborative Innovation Center for Green Transformation of Bio-resources, Hubei Key Laboratory of Industrial Biotechnology, College of Life Sciences, Hubei University, Wuhan, People's Republic of China.
authors.forename: Li
authors.initials: L
authors.lastname: Yi
authors.affiliation: Hubei Collaborative Innovation Center for Green Transformation of Bio-resources, Hubei Key Laboratory of Industrial Biotechnology, College of Life Sciences, Hubei University, Wuhan, People's Republic of China.
authors.forename: Lixin
authors.initials: L
authors.lastname: Ma
mesh_terms: D001119:Arginase / Q000235:genetics / Q000378:metabolism*; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000378:metabolism*; D005434:Flow Cytometry; D005822:Genetic Vectors / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008239:Lysine / Q000235:genetics / Q000378:metabolism; D008856:Microscopy, Fluorescence; D009842:Oligopeptides / Q000235:genetics / Q000378:metabolism; D009952:Ornithine / Q000378:metabolism; D015202:Protein Engineering; D021382:Protein Sorting Signals / Q000235:genetics; D014178:Translocation, Genetic
chemical_list: D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D009842:Oligopeptides; D021382:Protein Sorting Signals; C063113:ice nucleation protein; D009952:Ornithine; C574619:ARG1 protein, human; D001119:Arginase; D008239:Lysine