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Construction and characterization of β‐lactoglobulin chimeras
Author(s) -
Kobayashi Takuji,
Ikeguchi Masamichi,
Sugai Shintaro
Publication year - 2002
Publication title -
proteins: structure, function, and bioinformatics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.699
H-Index - 191
eISSN - 1097-0134
pISSN - 0887-3585
DOI - 10.1002/prot.10223
Subject(s) - mutant , dimer , chemistry , monomer , mutation , ultracentrifuge , mutant protein , microbiology and biotechnology , biology , biochemistry , gene , organic chemistry , polymer
At neutral pH, equine β‐lactoglobulin (ELG) is monomeric, whereas bovine β‐lactoglobulin (BLG) exists as a dimer. To understand the difference in the oligomerization properties between ELG and BLG, three mutants of ELG (LP, I, and LPI) were constructed by substituting amino acids responsible for important interactions at the dimer interface of BLG into ELG. The mutant LP has an AB loop mutation (S34A/E35Q), the mutant I has an I strand mutation (G145M/R146H/V147I/Q148R/I149L/V150S/P151F/D152N/L153P) and the mutant LPI includes both the LP and I mutations. The far‐ and near‐UV CD spectra of the three mutants are similar to that of the wild‐type ELG, indicating that the secondary and the tertiary structures of ELG are not significantly affected by the mutations. Ultracentrifuge analysis shows that all three mutants are monomeric at neutral pH, suggesting that the protein sequences in the AB loop and I strand of BLG alone cannot support dimerization of ELG. Thus, structural differences must exist between ELG and BLG that prevent the ELG mutants from forming the same interactions as BLG at the dimer interface. Proteins 2002;49:297–301. © 2002 Wiley‐Liss, Inc.