1. Intramolecular signal transmission in enterobacterial aspartate transcarbamylases II. Engineering co-operativity and allosteric regulation in the aspartate transcarbamylase of Erwinia herbicola.
- Author
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Cunin R, Rani CS, Van Vliet F, Wild JR, and Wales M
- Subjects
- Adenosine Triphosphate metabolism, Adenosine Triphosphate pharmacology, Allosteric Regulation drug effects, Amino Acid Sequence, Amino Acid Substitution, Aspartate Carbamoyltransferase antagonists & inhibitors, Aspartate Carbamoyltransferase chemistry, Aspartate Carbamoyltransferase genetics, Aspartic Acid metabolism, Binding, Competitive, Catalytic Domain, Cytidine Triphosphate antagonists & inhibitors, Cytidine Triphosphate metabolism, Cytidine Triphosphate pharmacology, Enterobacteriaceae genetics, Enzyme Activation drug effects, Escherichia coli genetics, Escherichia coli Proteins, Kinetics, Models, Molecular, Molecular Sequence Data, Recombinant Fusion Proteins antagonists & inhibitors, Recombinant Fusion Proteins chemistry, Recombinant Fusion Proteins genetics, Recombinant Fusion Proteins metabolism, Sequence Alignment, Structure-Activity Relationship, Uridine Triphosphate pharmacology, Aspartate Carbamoyltransferase metabolism, Enterobacteriaceae enzymology, Escherichia coli enzymology, Protein Engineering, Signal Transduction
- Abstract
The aspartate transcarbamylase (ATCase) from Erwinia herbicola differs from the other investigated enterobacterial ATCases by its absence of homotropic co-operativity toward the substrate aspartate and its lack of response to ATP which is an allosteric effector (activator) of this family of enzymes. Nevertheless, the E. herbicola ATCase has the same quaternary structure, two trimers of catalytic chains with three dimers of regulatory chains ((c3)2(r2)3), as other enterobacterial ATCases and shows extensive primary structure conservation. In (c3)2(r2)3 ATCases, the association of the catalytic subunits c3 with the regulatory subunits r2 is responsible for the establishment of positive co-operativity between catalytic sites for the binding of aspartate and it dictates the pattern of allosteric response toward nucleotide effectors. Alignment of the primary sequence of the regulatory polypeptides from the E. herbicola and from the paradigmatic Escherichia coli ATCases reveals major blocks of divergence, corresponding to discrete structural elements in the E. coli enzyme. Chimeric ATCases were constructed by exchanging these blocks of divergent sequence between these two ATCases. It was found that the amino acid composition of the outermost beta-strand of a five-stranded beta-sheet in the effector-binding domain of the regulatory polypeptide is responsible for the lack of co-operativity and response to ATP of the E. herbicola ATCase. A novel structural element involved in allosteric signal recognition and transmission in this family of ATCases was thus identified., (Copyright 1999 Academic Press.)
- Published
- 1999
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