NDLI: The 80s loop of the catalytic chain of Escherichia coli aspartate transcarbamoylase is critical for catalysis and homotropic cooperativity.

Content Provider	PubMed Central
Author	Macol, C. Dutta, M. Stec, B. Tsuruta, H. Kantrowitz, E. R.
Abstract	The X-ray structure of the Escherichia coli aspartate transcarbamoylase with the bisubstrate analog phosphonacetyl-L-aspartate (PALA) bound shows that PALA interacts with Lys84 from an adjacent catalytic chain. To probe the function of Lys84, site-specific mutagenesis was used to convert Lys84 to alanine, threonine, and asparagine. The K84N and K84T enzymes exhibited 0.08 and 0.29% of the activity of the wild-type enzyme, respectively. However, the K84A enzyme retained 12% of the activity of the wild-type enzyme. For each of these enzymes, the affinity for aspartate was reduced 5- to 10-fold, and the affinity for carbamoyl phosphate was reduced 10- to 30-fold. The enzymes K84N and K84T exhibited no appreciable cooperativity, whereas the K84A enzyme exhibited a Hill coefficient of 1.8. The residual cooperativity and enhanced activity of the K84A enzyme suggest that in this enzyme another mechanism functions to restore catalytic activity. Modeling studies as well as molecular dynamics simulations suggest that in the case of only the K84A enzyme, the lysine residue at position 83 can reorient into the active site and complement for the loss of Lys84. This hypothesis was tested by the creation and analysis of the K83A enzyme and a double mutant enzyme (DM) that has both Lys83 and Lys84 replaced by alanine. The DM enzyme has no cooperativity and exhibited 0.18% of wild-type activity, while the K83A enzyme exhibited 61% of wild-type activity. These data suggest that Lys84 is not only catalytically important, but is also essential for binding both substrates and creation of the high-activity, high-affinity active site. Since low-angle X-ray scattering demonstrated that the mutant enzymes can be converted to the R-structural state, the loss of cooperativity must be related to the inability of these mutant enzymes to form the high-activity, high-affinity active site characteristic of the R-functional state of the enzyme.
Related Links	http://dx.doi.org/10.1110/ps.8.6.1305
Ending Page	1313
Page Count	9
Starting Page	1305
File Format	PDF
ISSN	09618368
e-ISSN	1469896X
Journal	Protein Science : A Publication of the Protein Society
Issue Number	6
Volume Number	8
Language	English
Publisher	Cold Spring Harbor Laboratory Press
Publisher Date	1999-01-01
Access Restriction	Open
Rights Holder	Cold Spring Harbor Laboratory Press
Subject Keyword	Biochemistry Molecular Biology Research in Higher Education
Content Type	Text
Resource Type	Article
Subject	Medicine Molecular Biology Biochemistry

Sl.	Authority	Responsibilities	Communication Details
1	Ministry of Education (GoI), Department of Higher Education	Sanctioning Authority	https://www.education.gov.in/ict-initiatives
2	Indian Institute of Technology Kharagpur	Host Institute of the Project: The host institute of the project is responsible for providing infrastructure support and hosting the project	https://www.iitkgp.ac.in
3	National Digital Library of India Office, Indian Institute of Technology Kharagpur	The administrative and infrastructural headquarters of the project	Dr. B. Sutradhar bsutra@ndl.gov.in
4	Project PI / Joint PI	Principal Investigator and Joint Principal Investigators of the project	Dr. B. Sutradhar bsutra@ndl.gov.in Prof. Saswat Chakrabarti will be added soon
5	Website/Portal (Helpdesk)	Queries regarding NDLI and its services	support@ndl.gov.in
6	Contents and Copyright Issues	Queries related to content curation and copyright issues	content@ndl.gov.in
7	National Digital Library of India Club (NDLI Club)	Queries related to NDLI Club formation, support, user awareness program, seminar/symposium, collaboration, social media, promotion, and outreach	clubsupport@ndl.gov.in
8	Digital Preservation Centre (DPC)	Assistance with digitizing and archiving copyright-free printed books	dpc@ndl.gov.in
9	IDR Setup or Support	Queries related to establishment and support of Institutional Digital Repository (IDR) and IDR workshops	idr@ndl.gov.in

The 80s loop of the catalytic chain of Escherichia coli aspartate transcarbamoylase is critical for catalysis and homotropic cooperativity.

The importance of the link between Glu204 of the catalytic chain and Arg130 of the regulatory chain for the homotropic and heterotropic properties of Escherichia coli aspartate transcarbamoylase.

The regulatory subunit of Escherichia coli aspartate carbamoyltransferase may influence homotropic cooperativity and heterotropic interactions by a direct interaction with the loop containing residues 230-245 of the catalytic chain.

Allostery and cooperativity in Escherichia coli Aspartate Transcarbamoylase

Arginine 54 in the active site of Escherichia coli aspartate transcarbamoylase is critical for catalysis: a site-specific mutagenesis, NMR, and X-ray crystallographic study.

Glu-50 in the catalytic chain of Escherichia coli aspartate transcarbamoylase plays a crucial role in the stability of the R quaternary structure.

240s Loop Interactions Stabilize the T State of Escherichia coli Aspartate Transcarbamoylase

Weakening of the interface between adjacent catalytic chains promotes domain closure in Escherichia coli aspartate transcarbamoylase.

Stabilization of the R Allosteric Structure of Escherichia coli Aspartate Transcarbamoylase by Disulfide Bond Formation

The 80s loop of the catalytic chain of Escherichia coli aspartate transcarbamoylase is critical for catalysis and homotropic cooperativity.

Similar Documents

The 80s loop of the catalytic chain of Escherichia coli aspartate transcarbamoylase is critical for catalysis and homotropic cooperativity.

The importance of the link between Glu204 of the catalytic chain and Arg130 of the regulatory chain for the homotropic and heterotropic properties of Escherichia coli aspartate transcarbamoylase.

The regulatory subunit of Escherichia coli aspartate carbamoyltransferase may influence homotropic cooperativity and heterotropic interactions by a direct interaction with the loop containing residues 230-245 of the catalytic chain.

Allostery and cooperativity in Escherichia coli Aspartate Transcarbamoylase

Arginine 54 in the active site of Escherichia coli aspartate transcarbamoylase is critical for catalysis: a site-specific mutagenesis, NMR, and X-ray crystallographic study.

Glu-50 in the catalytic chain of Escherichia coli aspartate transcarbamoylase plays a crucial role in the stability of the R quaternary structure.

240s Loop Interactions Stabilize the T State of Escherichia coli Aspartate Transcarbamoylase

Weakening of the interface between adjacent catalytic chains promotes domain closure in Escherichia coli aspartate transcarbamoylase.

Stabilization of the R Allosteric Structure of Escherichia coli Aspartate Transcarbamoylase by Disulfide Bond Formation

The 80s loop of the catalytic chain of Escherichia coli aspartate transcarbamoylase is critical for catalysis and homotropic cooperativity.