Open AccessSurface Immobilization of Human Arginase-1 with an Engineered Ice Nucleation Protein Display System in E. coli
Author(s) -
Zhen Zhang,
Rongxin Tang,
Boxun Lu,
Meng Mei,
Chunhua Li,
Xinrong Ma,
Yang Li,
Lixin Ma
Publication year - 2016
Publication title -
plos one
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.99
H-Index - 332
ISSN - 1932-6203
DOI - 10.1371/journal.pone.0160367
Subject(s) - arginase , peptide , chemistry , arginine , nucleation , biochemistry , biophysics , amino acid , biology , organic chemistry
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 K 6 -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 K 6 -InaK-N/ARG1 presented an arginase activity of 10.7 U/OD 600 under the optimized conditions of 40°C, pH 10.0 and 1 mM Mn 2+ , 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.