Sequential truncation of the lactose permease over a three-amino acid sequence near the carboxyl terminus leads to progressive loss of activity and stability

Journal Article · Mon Apr 15 00:00:00 EDT 1991 · Proceedings of the National Academy of Sciences of the United States of America; (United States)

DOI:https://doi.org/10.1073/pnas.88.8.2969· OSTI ID:5934351

McKenna, E; Hardy, D; Pastore, J C; Kaback, H R ^[1]

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1. Univ. of California, Los Angeles (United States)

Previous experiments are consistent with the notion that residues 396-401 (...SVFTLS...) at the carboxyl terminus of the last putative transmembrane helix of the lactose (lac) permease of Escherichia coli are important for protection against proteolytic degradation and suggest that this region of the permease may be necessary for proper folding. Stop codons (TAA) have now been substituted sequentially for amino acid codons 396-401 in the lacY gene, and the termination mutants were expressed from the plasmid pT7-5. With respect to transport, permease truncated at residue 396-or 397 is completely defective, while molecules truncated at residues 398, 399, 400, and 401, respectively, exhibit 15-25%, 30-40%, 40-45%, and 70-100% of wild-type activity. As judged by pulse-chase experiments with ({sup 35}S)methionine, wild-type permease or permease truncated at residue 401 is stable, while permease molecules truncated at position 400, 399, 398, 397, or 396 are degraded at increasingly rapid rates. The findings indicate that either the last turn of putative helix XII or the region immediately distal to helix XII is important for proper folding and protection against proteolytic degradation.

• OSTI ID: 5934351
• Journal Information: Proceedings of the National Academy of Sciences of the United States of America; (United States), Vol. 88:8; ISSN 0027-8424
• Country of Publication: United States
• Language: English

Related Subjects

59 BASIC BIOLOGICAL SCIENCES
LACTOSE
MEMBRANE TRANSPORT
TRANSFERASES
AMINO ACID SEQUENCE
CARBON 14 COMPOUNDS
ESCHERICHIA COLI
LABELLED COMPOUNDS
MEMBRANE PROTEINS
METHIONINE
SULFUR 35
TRACER TECHNIQUES
AMINO ACIDS
BACTERIA
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
CARBOHYDRATES
CARBON COMPOUNDS
CARBOXYLIC ACIDS
DAYS LIVING RADIOISOTOPES
DISACCHARIDES
DRUGS
ENZYMES
EVEN-ODD NUCLEI
ISOTOPE APPLICATIONS
ISOTOPES
LIGHT NUCLEI
LIPOTROPIC FACTORS
MICROORGANISMS
MOLECULAR STRUCTURE
NUCLEI
OLIGOSACCHARIDES
ORGANIC ACIDS
ORGANIC COMPOUNDS
ORGANIC SULFUR COMPOUNDS
PROTEINS
RADIOISOTOPES
SACCHARIDES
SULFUR ISOTOPES
550201* - Biochemistry- Tracer Techniques