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Title 

Active human ferritin H/L-hybrid and sequence effect on folding efficiency in Escherichia coli

Authors 

Jee Won LeeSung Woo KimY H KimJ Y Ahn

Publisher 

Elsevier

Issue Date 

2002

Citation 

Biochemical and Biophysical Research Communications, vol. 298, no. 2, pp. 225-229

Keywords 

cytoplasmic solubilityferritin H/L-hybridfolding efficiencyintra-molecular interactioniron storage capacityceruloplasminferritinpolypeptideamino acid sequenceamino terminal sequence

Abstract 

In Escherichia coli, the recombinant human L-chain ferritin was synthesized in the form of inclusion bodies under the control of T7 promoter system. We developed a recombinant ferritin H/L-hybrid by a direct gene fusion between H- and L-chain subunits. Surprisingly, the presence of heavy-chain polypeptide at the amino terminus of light chain significantly increased the cytoplasmic solubility of the recombinant ferritin hybrid, i.e., more than 80% of synthesized ferritin hybrid was soluble in intracellular region regardless of the changes in cell growth and gene expression conditions such as type of inducer, growth media, culture scale, etc. The soluble ferritin H/L-hybrid was biologically active with the iron storage capacity (295 mol Fe+3 per mol H/L-hybrid) equivalent to ferritin standard. Different types of hybrid mutants were also developed using various H-chain derivatives. Comparison of the intracellular solubilities of the synthesized hybrid mutants showed that the N-terminus four helices of heavy subunit were of crucial importance in maintaining the high solubility in E. coli cytoplasm. Consequently, the increased solubility of the ferritin hybrid seems to be related to such H-chain sequence that forms ferroxidase center and promotes effective intra-molecular interaction with L-chain domain of H/L-hybrid for enhancing the folding efficiency.

ISSN 

0006-291X

Link 

http://dx.doi.org/10.1016/S0006-291X(02)02429-4

Appears in Collections

1. Journal Articles > Journal Articles

Registered Date

2019-05-02


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