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Engineered β-Lactoglobulin Produced in E. coli: Purification, Biophysical and Structural Characterisation.

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Loch JI, Bonarek P, Tworzydło M, Polit A, Hawro B, Łach A, Ludwin E, Lewiński K
OpenAccess logo Mol Biotechnol 58 605-618 (2016)
Related entries: 5htd, 5k06

Cited: 3 times
EuropePMC logo PMID: 27380951

Abstract

Functional recombinant bovine β-lactoglobulin has been produced by expression in E. coli using an engineered protein gene and purified to homogeneity by applying a new protocol. Mutations L1A/I2S introduced into the protein sequence greatly facilitate in vivo cleavage of the N-terminal methionine, allowing correctly folded and soluble protein suitable for biochemical, biophysical and structural studies to be obtained. The use of gel filtration on Sephadex G75 at the last purification step enables protein without endogenous ligand to be obtained. The physicochemical properties of recombinant β-lactoglobulin such as CD spectra, ligand binding (n, K a, ΔH, TΔS, ΔG), chemical and thermal stability (ΔG D, C mid) and crystal structure confirmed that the protein obtained is almost identical to the natural one. The substitutions of N-terminal residues did not influence the binding properties of the recombinant protein so that the lactoglobulin produced and purified according to our protocol is a good candidate for further engineering and potential use in pharmacology and medicine.

Articles - 5hte mentioned but not cited (1)

  1. Engineered β-Lactoglobulin Produced in E. coli: Purification, Biophysical and Structural Characterisation. Loch JI, Bonarek P, Tworzydło M, Polit A, Hawro B, Łach A, Ludwin E, Lewiński K. Mol Biotechnol 58 605-618 (2016)


Articles citing this publication (2)

  1. Methyl-selective isotope labeling using α-ketoisovalerate for the yeast Pichia pastoris recombinant protein expression system. Suzuki R, Sakakura M, Mori M, Fujii M, Akashi S, Takahashi H. J Biomol NMR 71 213-223 (2018)
  2. New ligand-binding sites identified in the crystal structures of β-lactoglobulin complexes with desipramine. Loch JI, Barciszewski J, Śliwiak J, Bonarek P, Wróbel P, Pokrywka K, Shabalin IG, Minor W, Jaskolski M, Lewiński K. IUCrJ 9 386-398 (2022)


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