Cloning of the surface layer gene sllB from Bacillus sphaericus ATCC 14577 and its heterologous expression and purification

Abstract

A cDNA fragment encoding the S-layer protein SllB cloned from Bacillus sphaericus ATCC 14577 was expressed on the surface of E. coli BL21 (DE3) cells and confirmed by the square lattice structure at the nanoscale level. The amplified gene fragment designed with PCR primers from a specified reference sequence (GenBank accession no. AJ849550) showed a high degree of sequence identity with the known sequences for S-layer protein. The best alignment scores were seen in B. sphaericus strains JG-A12 and NCTC9602, which code for a pre-form protein with a predicted cleavage site located between the two alanine residues 31 and 32. After this signal peptide sequence was removed, the mature protein had a molecular mass of 116.2613 kDa and a theoretical pI of 5.40. Further bioinformatic analysis revealed three S-layer homology (SLH) domains in the N-terminus of the mature protein, positioned at the 1–61, 63–128 and 137–197 residues. The mature S-layer protein was composed of alpha helices (24.86%), extended strands (27.01%), and rich random coils (48.13%). Bioinformatics-driven characterization of SllB may provide scientific evidence for further application of this gene in the fields of nanobiotechnology and biomimetics in the future.

Genome comparison provides molecular insights into the phylogeny of the reassigned new genus Lysinibacillus

“…Co/Zn/Cd/Mg/Ni cation transporters (6 genes), metal-dependent hydrolases (3 genes), membrane proteins related to metalloendopeptidases (3 genes), Zinc metalloprotease, Mn-containing catalase, Fe-S cluster formation, and other related Oxidoreductases. KTCC 3346 contained 19 unique genes related to cell wall/membrane/envelope biogenesis, which may be associated with its ability to produce specific surface layer proteins [ 18 - 20 ]. It was also interesting that a gene homologous to the virion core protein of lumpy skin disease virus was identified in the marine strain Bacillus sp.…”