Structural role for Tyr-104 in Escherichia coli isopentenyl-diphosphate isomerase: site-directed mutagenesis, enzymology, and protein crystallography.
Isopentenyl-diphosphate (IPP):dimethylallyl diphosphate isomerase is a key enzyme in the biosynthesis of isoprenoids. The mechanism of the isomerization reaction involves protonation of the unactivated carbon-carbon double bond in the substrate, but identity of the acidic moiety providing the proton is still not clear. Multiple sequence alignments and geometrical features observed in crystal structures of complexes with IPP isomerase suggest that Tyr-104 could play an important role during catalysis. A series of mutants was constructed by directed mutagenesis and characterized by enzymology. Crystallographic and thermal denaturation data for Y104A and Y104F mutants were obtained. Those data demonstrate the importance of residue Tyr-104 for proper folding of Escherichia coli type I IPP isomerase.
de Ruyck
Jérôme
J
Laboratoire de Chimie Biologique Structurale, University of Namur, 61 Rue de Bruxelles, 5000 Namur, Belgium. jerome.deruyck@fundp.ac.be
Durisotti
Virginie
V
Oudjama
Yamina
Y
Wouters
Johan
J
D019748:Carbon-Carbon Double Bond Isomerases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D002384:Catalysis; D003461:Crystallography; D004926:Escherichia coli / Q000201:enzymology*; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D045782:Hemiterpenes; D016297:Mutagenesis, Site-Directed; D011489:Protein Denaturation; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D013329:Structure-Activity Relationship; D014443:Tyrosine / Q000378:metabolism
D029968:Escherichia coli Proteins; D045782:Hemiterpenes; D014443:Tyrosine; D019748:Carbon-Carbon Double Bond Isomerases; C021453:isopentenyldiphosphate delta-isomerase