scholarly journals Modulation of the ligand binding properties of the transcription repressor NmrA by GATA-containing DNA and site-directed mutagenesis

2009 ◽  
Vol 13 (12) ◽  
pp. 3127-3138 ◽  
Author(s):  
Heather K. Lamb ◽  
Jingshan Ren ◽  
Alison Park ◽  
Christopher Johnson ◽  
Kris Leslie ◽  
...  
Keyword(s):  
Ligand Binding ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Binding Properties

Role of Cysteine Residues in the N-Terminal Extracellular Domain of the Human VIP 1 Receptor for Ligand Binding A Site-directed Mutagenesis Studya

2006 ◽  
Vol 805 (1) ◽  
pp. 585-589 ◽  
Author(s):  
PASCALE GAUDIN ◽  
ALAIN COUVINEAU ◽  
JEAN-JOSÉ MAORET ◽  
CHRISTIANE ROUYER-FESSARD ◽  
MARC LABURTHE
Keyword(s):  
Ligand Binding ◽  
Extracellular Domain ◽  
Site Directed Mutagenesis ◽  
Cysteine Residues ◽  
Directed Mutagenesis ◽  
A Site

scholarly journals The role of Val68(E11) in ligand binding to sperm whale myoglobin. Site-directed mutagenesis of a synthetic gene.

1990 ◽  
Vol 265 (20) ◽  
pp. 11788-11795
Author(s):  
K D Egeberg ◽  
B A Springer ◽  
S G Sligar ◽  
T E Carver ◽  
R J Rohlfs ◽  
...  
Keyword(s):  
Ligand Binding ◽  
Sperm Whale ◽  
Site Directed Mutagenesis ◽  
Synthetic Gene ◽  
Directed Mutagenesis ◽  
Sperm Whale Myoglobin

scholarly journals Identification by site-directed mutagenesis of amino acids contributing to ligand-binding specificity or signal transduction properties of the human FP prostanoid receptor

2003 ◽  
Vol 371 (2) ◽  
pp. 443-449 ◽  
Author(s):  
Frank NEUSCHÄFER-RUBE ◽  
Eva ENGEMAIER ◽  
Sina KOCH ◽  
Ulrike BÖER ◽  
Gerhard P. PÜSCHEL
Keyword(s):  
Amino Acids ◽  
Ligand Binding ◽  
G Protein ◽  
Transmembrane Domain ◽  
Sequence Similarity ◽  
Membrane Receptors ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Receptor Subtypes ◽  
Plasma Membrane Receptors

Prostanoid receptors belong to the class of heptahelical plasma membrane receptors. For the five prostanoids, eight receptor subtypes have been identified. They display an overall sequence similarity of roughly 30%. Based on sequence comparison, single amino acids in different subtypes of different species have previously been identified by site-directed mutagenesis or in hybrid receptors that appear to be essential for ligand binding or G-protein coupling. Based on this information, a series of mutants of the human FP receptor was generated and characterized in ligand-binding and second-messenger-formation studies. It was found that mutation of His-81 to Ala in transmembrane domain 2 and of Arg-291 to Leu in transmembrane domain 7, which are putative interaction partners for the prostanoid's carboxyl group, abolished ligand binding. Mutants in which Ser-263 in transmembrane domain 6 or Asp-300 in transmembrane domain 7 had been replaced by Ala or Gln, respectively, no longer discriminated between prostaglandins PGF2α and PGD2. Thus distortion of the topology of transmembrane domains 6 and 7 appears to interfere with the cyclopentane ring selectivity of the receptor. PGF2α-induced inositol formation was strongly reduced in the mutant Asp-300Gln, inferring a role for this residue in agonist-induced G-protein activation.

scholarly journals The mechanism of action of dd-peptidases: the role of Threonine-299 and -301 in the Streptomyces R61 dd-peptidase

1994 ◽  
Vol 301 (2) ◽  
pp. 477-483 ◽  
Author(s):  
J M Wilkin ◽  
A Dubus ◽  
B Joris ◽  
J M Frère
Keyword(s):  
Amino Acids ◽  
Active Site ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Peptide Substrate ◽  
Binding Properties ◽  
Beta Lactamases ◽  
Penicillin Binding Proteins ◽  
Active Site Cavity

The side chains of residues Thr299 and Thr301 in the Streptomyces R61 DD-peptidase have been modified by site-directed mutagenesis. These amino acids are part of a beta-strand which forms a wall of the active-site cavity. Thr299 corresponds to the second residue of the Lys-Thr(Ser)-Gly triad, highly conserved in active-site beta-lactamases and penicillin-binding proteins (PBPs). Modification of Thr301 resulted only in minor alterations of the catalytic and penicillin-binding properties of the enzyme. No selective decrease of the rate of acylation was observed for any particular class of compounds. By contrast, the loss of the hydroxy group of the residue in position 299 yielded a seriously impaired enzyme. The rates of inactivation by penicillins were decreased 30-50-fold, whereas the reactions with cephalosporins were even more affected. The efficiency of hydrolysis against the peptide substrate was also seriously decreased. More surprisingly, the mutant was completely unable to catalyse transpeptidation reactions. The conservation of an hydroxylated residue in this position in PBPs is thus easily explained by these results.

scholarly journals Expression and site-directed mutagenesis of mouse prostaglandin E2 receptor EP3 subtype in insect cells

1995 ◽  
Vol 307 (2) ◽  
pp. 493-498 ◽  
Author(s):  
C Huang ◽  
H H Tai
Keyword(s):  
Ligand Binding ◽  
Prostaglandin E2 ◽  
Insect Cells ◽  
Expression System ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Wild Type ◽  
Binding Studies ◽  
Comparable Amount ◽  
Ep3 Receptor

A cDNA encoding for mouse prostaglandin E2 (PGE2) receptor EP3 subtype was cloned from a mouse kidney cDNA library by PCR using terminal primers derived from the known sequence of mouse lung EP3 receptor cDNA. The cloned cDNA was confirmed by sequencing and was expressed in Trichoplusia ni (MG1) insect cells using a baculovirus expression system. A specific protein of 60 kDa was detected by immunoblot with antibodies generated against a unique decapeptide sequence present in the second extracellular loop of the EP3 receptor. Specific binding of [3H]PGE2 with a Kd of 3 nM was also found in the membrane fraction of the insect cells. Ligand binding of the receptor was further studied by site-directed mutagenesis. Arg-309 of the receptor was separately mutated to lysine, glutamate and valine. cDNAs of the wild-type and mutant EP3 receptors were respectively expressed and studied in MG1 insect cells. Binding studies indicated that both glutamate and valine mutant EP3 receptors had no binding of [3H]PGE2. On the contrary, the lysine mutant receptor exhibited an even tighter binding (Kd = 1.3 nM) than the wild-type EP3 receptor. Immunoblot studies indicated that these receptors were expressed in a comparable amount in MG1 insect cells. These results suggest that Arg-309 of EP3 receptor may be essential in ligand binding through ionic interaction.

scholarly journals Analysis of ligand binding to a ribose biosensor using site-directed mutagenesis and fluorescence spectroscopy

2007 ◽  
Vol 16 (3) ◽  
pp. 362-368 ◽  
Author(s):  
N. C. Vercillo ◽  
K. J. Herald ◽  
J. M. Fox ◽  
B. S. Der ◽  
J. D. Dattelbaum
Keyword(s):  
Fluorescence Spectroscopy ◽  
Ligand Binding ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis
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