Glutamine-dependent NAD+ synthetase. How a two-domain, three-substrate enzyme avoids waste

Marzena Wojcik; Heather F Seidle; Pawel Bieganowski; Charles Brenner

doi:10.1074/jbc.M607111200

Back

Glutamine-dependent NAD+ synthetase. How a two-domain, three-substrate enzyme avoids waste

Journal article

Open access

Peer reviewed

Glutamine-dependent NAD+ synthetase. How a two-domain, three-substrate enzyme avoids waste

Marzena Wojcik, Heather F Seidle, Pawel Bieganowski and Charles Brenner

The Journal of biological chemistry, Vol.281(44), pp.33395-33402

11/03/2006

DOI: 10.1074/jbc.M607111200

PMID: 16954203

Files and links (1)

url

https://doi.org/10.1074/jbc.M607111200View

Published (Version of record) Open Access

Abstract

Glutamine-dependent NAD(+) synthetase, Qns1, utilizes a glutamine aminotransferase domain to supply ammonia for amidation of nicotinic acid adenine dinucleotide (NaAD(+)) to NAD(+). Earlier characterization of Qns1 suggested that glutamine consumption exceeds NAD(+) production by 40%. To explore whether Qns1 is systematically wasteful or whether additional features account for this behavior, we performed a careful kinetic and molecular genetic analysis. In fact, Qns1 possesses remarkable properties to reduce waste. The glutaminase active site is stimulated by NaAD(+) more than 50-fold such that glutamine is not appreciably consumed in the absence of NaAD(+). Glutamine consumption exceeds NAD(+) production over the whole range of glutamine and NaAD(+) substrate concentrations with greatest efficiency occurring at saturation of both substrates. Kinetic data coupled with site-directed mutagenesis of amino acids in the predicted ammonia channel indicate that NaAD(+) stimulates the glutaminase active site in the k(cat) term by a synergistic mechanism that does not require ammonia utilization by the NaAD(+) substrate. Six distinct classes of Qns1 mutants that fall within the glutaminase domain and the synthetase domain selectively inhibit components of the coordinated reaction.

Adenosine Triphosphate - pharmacology

NAD - analogs & derivatives

Ammonia - metabolism

Models, Biological

Glutamine - metabolism

Substrate Specificity

Kinetics

Mutation - genetics

Amide Synthases - classification

Binding Sites

NAD - metabolism

Amide Synthases - metabolism

Details

Title: Subtitle: Glutamine-dependent NAD+ synthetase. How a two-domain, three-substrate enzyme avoids waste
Creators: Marzena Wojcik - Department of Genetics and the Norris Cotton Cancer Center, Dartmouth Medical School, Lebanon, New Hampshire 03756, USA
Heather F Seidle
Pawel Bieganowski
Charles Brenner
Resource Type: Journal article
Publication Details: The Journal of biological chemistry, Vol.281(44), pp.33395-33402
DOI: 10.1074/jbc.M607111200
PMID: 16954203
NLM abbreviation: J Biol Chem
ISSN: 0021-9258
eISSN: 1083-351X
Publisher: United States
Grant note: P01 CA077738-01A20003 / NCI NIH HHS P01 CA077738 / NCI NIH HHS
Language: English
Date published: 11/03/2006
Academic Unit: Biochemistry and Molecular Biology; Internal Medicine
Record Identifier: 9983788598802771

Metrics

33 Record Views

46 Times Cited - Web of Science