The active site and mechanism of the β-galactosidase from Escherichia coli

Huber, R.E.; Gupta, Munishwar N.; Khare, Sunil Kumar

doi:10.1016/0020-711x(94)90051-5

Cited by 49 publications

(21 citation statements)

References 94 publications

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“…4 gives the b-galactosidase mediated gain in allolactose from the conversion of lactose following the studies of Huber et al (1976). The second term accounts for allolactose loss via conversion to glucose and galactose, again mediated by b-galactosidase (Martínez-Bilbao et al, 1991;Huber et al, 1994). The last term takes into account the degradation and dilution of allolactose.…”

Section: The Modelmentioning

confidence: 99%

Feedback Regulation in the Lactose Operon: A Mathematical Modeling Study and Comparison with Experimental Data

Yıldırım

Mackey

2003

Biophysical Journal

193

233

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A mathematical model for the regulation of induction in the lac operon in Escherichia coli is presented. This model takes into account the dynamics of the permease facilitating the internalization of external lactose; internal lactose; beta-galactosidase, which is involved in the conversion of lactose to allolactose, glucose and galactose; the allolactose interactions with the lac repressor; and mRNA. The final model consists of five nonlinear differential delay equations with delays due to the transcription and translation process. We have paid particular attention to the estimation of the parameters in the model. We have tested our model against two sets of beta-galactosidase activity versus time data, as well as a set of data on beta-galactosidase activity during periodic phosphate feeding. In all three cases we find excellent agreement between the data and the model predictions. Analytical and numerical studies also indicate that for physiologically realistic values of the external lactose and the bacterial growth rate, a regime exists where there may be bistable steady-state behavior, and that this corresponds to a cusp bifurcation in the model dynamics.

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Section: The Modelmentioning

confidence: 99%

Feedback Regulation in the Lactose Operon: A Mathematical Modeling Study and Comparison with Experimental Data

Yıldırım

Mackey

2003

Biophysical Journal

193

233

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“…It catalyzes the cleavage of the bond between the anomeric carbon and glycosyl oxygen of a β-D-galactopyranoside [71]. In vivo LacZ catalyzes the cleavage of the disaccharide lactose to form glucose and galactose [72].…”

Section: Resultsmentioning

confidence: 99%

PDZ affinity chromatography: A general method for affinity purification of proteins based on PDZ domains and their ligands

Walkup

Kennedy

2014

Protein Expression and Purification

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PDZ (PSD-95, DiscsLarge, ZO1) domains function in nature as protein binding domains within scaffold and membrane-associated proteins. They comprise ~ 90 residues and make specific, high affinity interactions with complementary C-terminal peptide sequences, with other PDZ domains, and with phospholipids. We hypothesized that the specific, strong interactions of PDZ domains with their ligands would make them well suited for use in affinity chromatography. Here we describe a novel affinity chromatography method applicable for the purification of proteins that contain PDZ domain-binding ligands, either naturally or introduced by genetic engineering. We created a series of affinity resins comprised of PDZ domains from the scaffold protein PSD-95, or from neuronal nitric oxide synthase (nNOS), coupled to solid supports. We used them to purify heterologously expressed neuronal proteins or protein domains containing endogenous PDZ domain ligands, eluting the proteins with free PDZ domain peptide ligands. We show that Proteins of Interest (POIs) lacking endogenous PDZ domain ligands can be engineered as fusion products containing C-terminal PDZ domain ligand peptides or internal, N- or C-terminal PDZ domains and then can be purified by the same method. Using this method, we recovered recombinant GFP fused to a PDZ-domain ligand in active form as verified by fluorescence yield. Similarly, chloramphenicol acetyltransferase (CAT) and β-Galactosidase (LacZ) fused to a C-terminal PDZ domain ligand or an N-terminal PDZ domain were purified in active form as assessed by enzymatic assay. In general, PDZ domains and ligands derived from PSD-95 were superior to those from nNOS for this method. PDZ Domain Affinity Chromatography promises to be a versatile and effective method for purification of a wide variety of natural and recombinant proteins.

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“…The primary enzymatic function of ␤-gal relevant to its role as a biotechnological tool is to cleave the chemical bond between the anomeric carbon and glycosyl oxygen of appropriate substrates ( Fig. 1; see (13,14) for review). While cleaved substrates are restricted to ␤-D-galactopyranosides, the enzyme is tolerant of a range of aglycone groups of differing chemical composition.…”

Section: The Origins Of Laczmentioning

confidence: 99%

Uses of lacZ to Study Gene Function: Evaluation of β-Galactosidase Assays Employed in the Yeast Two-Hybrid System

Serebriiskii

Golemis

2000

Analytical Biochemistry

114

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The active site and mechanism of the β-galactosidase from Escherichia coli

Cited by 49 publications

References 94 publications

Feedback Regulation in the Lactose Operon: A Mathematical Modeling Study and Comparison with Experimental Data

Feedback Regulation in the Lactose Operon: A Mathematical Modeling Study and Comparison with Experimental Data

PDZ affinity chromatography: A general method for affinity purification of proteins based on PDZ domains and their ligands

Uses of lacZ to Study Gene Function: Evaluation of β-Galactosidase Assays Employed in the Yeast Two-Hybrid System

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