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Invitro complex‐formation between the molecular chaperone DnaK and staphylococcal ProteinA derivatives produced in Escherichia coli and its use inthe purification of DnaK
Author(s) -
Gustavsson Kristina,
Bergman Tomas,
Veide Andres,
Enfors SvenOlof
Publication year - 1997
Publication title -
biotechnology and applied biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.468
H-Index - 70
eISSN - 1470-8744
pISSN - 0885-4513
DOI - 10.1111/j.1470-8744.1997.tb00430.x
Subject(s) - proteolysis , escherichia coli , biochemistry , chaperone (clinical) , biology , tryptophan , in vitro , incubation , peptide , amino acid , enzyme , medicine , pathology , gene
Complex‐formation between a truncated staphylococcal Protein A produced in Escherichia coli and a native E. coli molecular chaperone, DnaK, can be used for the purification of DnaK by IgG‐affinity chromatography. The half‐time constant for in vitro formation of the Protein AŐDnaK complex is about 14 min. Complex‐formation in the presence of ATP is faster, but pre‐incubation of DnaK with ATP decreases the final amount of the complex. A second complex with a slower migration on native PAGE is formed when the ratio of DnaK to Protein A is increased. A derivative of Protein A, ZZ, which essentially contains only two modified domains of Protein A, did not bind DnaK. After insertion of a tryptophan‐rich peptide close to the C‐terminus, the resulting protein, ZZT 3 , became able to bind DnaK. The binding of these three proteins to DnaK correlates with proteolysis in E. coli , indicating a possible role for the binding of DnaK in the control of proteolysis.