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Human Dipeptidyl Peptidase III: Insights into Ligand Binding from a Combined Experimental and Computational Approach


Tomić, Antonija; Abramić, Marija; Špoljarić, Jasminka; Agić, Dejan; Smith, David M.; Tomić, Sanja
Human Dipeptidyl Peptidase III: Insights into Ligand Binding from a Combined Experimental and Computational Approach // Journal of molecular recognition, 24 (2011), 5; 804-814 doi:10.1002/jmr.1115 (međunarodna recenzija, članak, znanstveni)


Naslov
Human Dipeptidyl Peptidase III: Insights into Ligand Binding from a Combined Experimental and Computational Approach

Autori
Tomić, Antonija ; Abramić, Marija ; Špoljarić, Jasminka ; Agić, Dejan ; Smith, David M. ; Tomić, Sanja

Izvornik
Journal of molecular recognition (0952-3499) 24 (2011), 5; 804-814

Vrsta, podvrsta i kategorija rada
Radovi u časopisima, članak, znanstveni

Ključne riječi
Dipeptidyl peptidase III; molecular modelling; binding affinity; catalytic affinity
(Dipeptidyl peptidase III molecular modelling; binding affinity; catalytic affinity)

Sažetak
Human dipeptidyl peptidase III (DPP III) is a zinc-exopeptidase with implied roles in protein catabolism, pain modulation and defense against oxidative stress. To understand the mode of ligand binding into its active site, we performed molecular modeling, site-directed mutagenesis and biochemical analyses. Using the recently determined crystal structure of the human DPP III we built complexes between both, the wild type protein and its mutant H568N with the preferred substrate Arg-Arg-2-naphthylamide (RRNA) and a competitive inhibitor Tyr-Phe-hydroxamate (Tyr-Phe-NHOH). The mutation of the conserved H1099-1352is568, structurally equivalent to catalytically important His231 in thermolysin, to Asn, resulted in a 1300-fold decrease of kcat for RRNA hydrolysis and in significantly lowered affinity for the inhibitor. Molecular dynamics (MD) simulations revealed the key protein ligand interactions as well as the ligand induced reorganization of the binding site and its partial closure. Simultaneously, the non catalytic domain was observed to stretch and the opening at the wide side of the inter-domain cleft became enhanced. The driving force for these changes was the formation of the hydrogen bond between Asp372 and the bound ligand. The structural and dynamical differences, found for the ligand binding to the wild-type enzyme and the H568N mutant, and the calculated binding free energies, agree well with the measured affinities. On the basis of the obtained results we suggest a possible reaction mechanism. In addition, this work provides a foundation for further site-directed mutagenesis experiments, as well as for modeling the reaction itself.

Izvorni jezik
Engleski

Znanstvena područja
Fizika, Kemija, Biologija



POVEZANOST RADA


Projekt / tema
022-0222148-2822 - Modeliranje i međudjelovanje kompleksa prijelaznih metala i bioliganada (Jasmina Sabolović, )
098-0982933-2937 - Računalno proučavanje strukture i funkcije proteina (David Matthew Smith, )
098-1191344-2860 - Proučavanje biomakromolekula računalnim metodama i razvoj novih algoritama (Sanja Tomić, )
098-1191344-2938 - Molekularna enzimologija i proteinske interakcije hidrolaza (Marija Abramić, )

Ustanove
Institut za medicinska istraživanja i medicinu rada, Zagreb,
Institut "Ruđer Bošković", Zagreb

Časopis indeksira:


  • Current Contents Connect (CCC)
  • Web of Science Core Collection (WoSCC)
    • Science Citation Index Expanded (SCI-EXP)
    • SCI-EXP, SSCI i/ili A&HCI
  • Scopus
  • MEDLINE


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