Many prokaryotic soluble PPases (pyrophosphatases) contain a pair of regulatory adenine nucleotide-binding CBS (cystathionine β-synthase) domains that act as ‘internal inhibitors’ whose effect is modulated by nucleotide binding. Although such regulatory domains are found in important enzymes and transporters, the underlying regulatory mechanism has only begun to come into focus. We reported previously that CBS domains bind nucleotides co-operatively and induce positive kinetic co-operativity (non-Michaelian behaviour) in CBS-PPases (CBS domain-containing PPases). In the present study, we demonstrate that a homodimeric ehPPase (Ethanoligenens harbinense PPase) containing an inherent mutation in an otherwise conserved asparagine residue in a loop near the active site exhibits non-co-operative hydrolysis kinetics. A similar N312S substitution in ‘co-operative’ dhPPase (Desulfitobacterium hafniense PPase) abolished kinetic co-operativity while causing only minor effects on nucleotide-binding affinity and co-operativity. However, the substitution reversed the effect of diadenosine tetraphosphate, abolishing kinetic co-operativity in wild-type dhPPase, but restoring it in the variant dhPPase. A reverse serine-to-asparagine replacement restored kinetic co-operativity in ehPPase. Molecular dynamics simulations revealed that the asparagine substitution resulted in a change in the hydrogen-bonding pattern around the asparagine residue and the subunit interface, allowing greater flexibility at the subunit interface without a marked effect on the overall structure. These findings identify this asparagine residue as lying at the ‘crossroads’ of information paths connecting catalytic and regulatory domains within a subunit and catalytic sites between subunits.
Skip Nav Destination
Article navigation
July 2016
-
Cover Image
Cover Image
Bacterial heme transporter HasR with its signaling domain. For further information, please see pp. 2239–2248. Image kindly provided by Nadia Izadi-Pruneyre. - PDF Icon PDF LinkFront Matter
- PDF Icon PDF LinkTable of Contents
- PDF Icon PDF LinkEditorial Board
Research Article|
July 12 2016
An asparagine residue mediates intramolecular communication in nucleotide-regulated pyrophosphatase
Viktor A. Anashkin;
Viktor A. Anashkin
1
*Department of Biochemistry, University of Turku, FIN-20014 Turku, Finland
†Belozersky Institute of Physico-Chemical Biology and Department of Chemistry, Lomonosov Moscow State University, Moscow 119899, Russia
Search for other works by this author on:
Anu Salminen;
Anu Salminen
1
*Department of Biochemistry, University of Turku, FIN-20014 Turku, Finland
Search for other works by this author on:
Natalia N. Vorobjeva;
Natalia N. Vorobjeva
†Belozersky Institute of Physico-Chemical Biology and Department of Chemistry, Lomonosov Moscow State University, Moscow 119899, Russia
Search for other works by this author on:
Reijo Lahti;
Reijo Lahti
2
*Department of Biochemistry, University of Turku, FIN-20014 Turku, Finland
2Correspondence may be addressed to either of these authors (email reijo.lahti@utu.fi or baykov@genebee.msu.su).
Search for other works by this author on:
Alexander A. Baykov
Alexander A. Baykov
2
†Belozersky Institute of Physico-Chemical Biology and Department of Chemistry, Lomonosov Moscow State University, Moscow 119899, Russia
2Correspondence may be addressed to either of these authors (email reijo.lahti@utu.fi or baykov@genebee.msu.su).
Search for other works by this author on:
Publisher: Portland Press Ltd
Received:
April 05 2016
Revision Received:
May 11 2016
Accepted:
May 16 2016
Accepted Manuscript online:
May 17 2016
Online ISSN: 1470-8728
Print ISSN: 0264-6021
© 2016 The Author(s). published by Portland Press Limited on behalf of the Biochemical Society
2016
Biochem J (2016) 473 (14): 2097–2107.
Article history
Received:
April 05 2016
Revision Received:
May 11 2016
Accepted:
May 16 2016
Accepted Manuscript online:
May 17 2016
Citation
Viktor A. Anashkin, Anu Salminen, Natalia N. Vorobjeva, Reijo Lahti, Alexander A. Baykov; An asparagine residue mediates intramolecular communication in nucleotide-regulated pyrophosphatase. Biochem J 15 July 2016; 473 (14): 2097–2107. doi: https://doi.org/10.1042/BCJ20160293
Download citation file:
Sign in
Don't already have an account? Register
Sign in to your personal account
You could not be signed in. Please check your email address / username and password and try again.
Captcha Validation Error. Please try again.