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Fine tuning the N‐terminus of a calcium binding protein: α‐lactalbumin
Author(s) -
Veprintsev Dmitry B.,
Narayan Mahesh,
Permyakov Serge E.,
Uversky Vladimir N.,
Brooks Charles L.,
Cherskaya Alexandra M.,
Permyakov Eugene A.,
Berliner Lawrence J.
Publication year - 1999
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/(sici)1097-0134(19991001)37:1<65::aid-prot7>3.0.co;2-2
Subject(s) - thermostability , lactalbumin , circular dichroism , chemistry , calcium , biochemistry , recombinant dna , native state , molten globule , protein secondary structure , methionine , amino acid , wild type , mutant , enzyme , organic chemistry , gene
The effects of amino acid substitutions in the N‐terminus of bovine recombinant α‐lactalbumin (including enzymatic removal of the N‐terminal methionine and deletion of Glu‐1) were studied by intrinsic fluorescence, circular dichroism (CD), and differential scanning microcalorimetry (DSC). Wild‐type recombinant α‐lactalbumin has a lower thermostability and calcium affinity compared to the native protein, while the properties of wild‐type protein with the N‐terminal methionine enzymatically removed are similar to the native protein. Taken together, the fluorescence, CD, and DSC results show that recombinant wild type α‐lactalbumin in the absence of calcium ion is in a type of molten globule state. The delta‐E1 mutant, where the Glu 1 residue of the native sequence is genetically removed, leaving an N‐terminal methionine in its place, shows almost one order of magnitude higher affinity for calcium and higher thermostability (both in the absence and presence of calcium) than the native protein isolated from milk. It was concluded that the N‐terminus of the protein dramatically affects both stability and function as manifested in calcium affinity. Proteins 1999;37:65–72. © 1999 Wiley‐Liss, Inc.