TY - JOUR
T1 - Oligonucleotide-directed site-specific mutagenesis of the 1ac permease of Escherichia coli
AU - Sarkar, Hemanta K.
AU - Viitanen, Paul V.
AU - Padan, Etana
AU - Trumble, William R.
AU - Poonian, Mohindar S.
AU - McComas, Warren
AU - Ronald Kaback, H.
PY - 1986/1/1
Y1 - 1986/1/1
N2 - This chapter illustrates that with the recent advent of site-directed mutagenesis, the goal of determining the role of amino acid residues or sequences in the function of a particular protein or enzyme, can be realized. The chapter describes an application of the technique to the lac permease of Escherichia coli. The lac permease is an intrinsic membrane protein, encoded by the lacY gene, that catalyzes symport (cotransport) of β-galactosides with protons. The lacY gene has been cloned and sequenced, and the permease has been purified to homogeneity in a completely functional state. A putative secondary structure model has been proposed based on the circular dichroic spectrum of the purified protein and on the sequential hydropathic character of the amino acid sequence. Although chemical modification of specific amino acid residues in a protein can provide important information, there are drawbacks to this approach. For this reason, oligonucleotide-directed, sitespecific mutageneses to study the structure and function of the lac permease have been utilized.
AB - This chapter illustrates that with the recent advent of site-directed mutagenesis, the goal of determining the role of amino acid residues or sequences in the function of a particular protein or enzyme, can be realized. The chapter describes an application of the technique to the lac permease of Escherichia coli. The lac permease is an intrinsic membrane protein, encoded by the lacY gene, that catalyzes symport (cotransport) of β-galactosides with protons. The lacY gene has been cloned and sequenced, and the permease has been purified to homogeneity in a completely functional state. A putative secondary structure model has been proposed based on the circular dichroic spectrum of the purified protein and on the sequential hydropathic character of the amino acid sequence. Although chemical modification of specific amino acid residues in a protein can provide important information, there are drawbacks to this approach. For this reason, oligonucleotide-directed, sitespecific mutageneses to study the structure and function of the lac permease have been utilized.
UR - http://www.scopus.com/inward/record.url?scp=0022558542&partnerID=8YFLogxK
U2 - 10.1016/S0076-6879(86)25019-3
DO - 10.1016/S0076-6879(86)25019-3
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C2 - 3520222
AN - SCOPUS:0022558542
SN - 0076-6879
VL - 125
SP - 214
EP - 230
JO - Methods in Enzymology
JF - Methods in Enzymology
IS - C
ER -