A new class of point mutations in the lacI gene of Escherichia coli is described which results in a lac repressor that is unable to form its normal tetrameric structure. Many of these mutations result in amino acid substitutions in the neighborhood of the tyrosine at residue 269. An amber mutation derived from the codon normally specifying tyrosine 269 has been suppressed with Sul, which inserts serine in response to the amber codon. The resulting altered lac repressor is also unable to form its normal tetrameric structure and can no longer bind to the lac operator. Two missense mutations which map in the part of the gene specifying amino acid residues 210 to 216 also produce repressors unable to form tetramers.

Bacterial Proteins↗

PubMedDate not supplied

Recombinant-derived interleukin-1 alpha stabilized against specific deamidation.

Recombinant-derived human interleukin-1 alpha (IL-1 alpha), purified from Escherichia coli, was resolved by isoelectric focusing on polyacrylamide gels into two species of isoelectric points (pI) 5.45 and 5.20, which constituted approximately 75% and approximately 25% of the total IL-1 alpha protein respectively. The pI 5.45 and pI 5.20 species were separated by chromatofocusing and subjected to N-terminal sequence analysis. The pI 5.45 species contained the expected Asn residue at position 36 of the mature protein sequence whereas the pI 5.20 species contained an Asp residue at the same position. A mutant protein in which Asn-36 was substituted for a Ser residue was isolated from E. coli and shown to be homogeneous on isoelectric focusing analysis with a pI = 5.45. 1H-n.m.r. and circular dichroism analyses of wild-type and the mutant IL-1 alpha indicated a similar conformation which was also indicated by the identical receptor binding affinities of IL-1 alpha with Asn, Asp or Ser in position 36. The mutant protein was stabilized against specific base-catalysed and temperature-induced deamidation, and may be more suitable than the wild-type position for physical and structural studies.

Amino Acids↗