The calcium-binding site of human glutamate carboxypeptidase II is critical for dimerization, thermal stability, and enzymatic activity

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F61388963%3A_____%2F18%3A00495871" target="_blank" >RIV/61388963:_____/18:00495871 - isvavai.cz</a>

Nalezeny alternativní kódy

RIV/00216208:11310/18:10386439 RIV/86652036:_____/18:00495871

Výsledek na webu

<a href="https://onlinelibrary.wiley.com/doi/full/10.1002/pro.3460" target="_blank" >https://onlinelibrary.wiley.com/doi/full/10.1002/pro.3460</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.1002/pro.3460" target="_blank" >10.1002/pro.3460</a>

Jazyk výsledku

angličtina

Název v původním jazyce

The calcium-binding site of human glutamate carboxypeptidase II is critical for dimerization, thermal stability, and enzymatic activity

Popis výsledku v původním jazyce

Calcium ions are required for proper function of a wide spectrum of proteins within cells. X-ray crystallography of human glutamate carboxypeptidase II (GCPII) revealed the presence of a Ca2+-binding site, but its importance for the structure and function of this metallopeptidase has not been elucidated to date. Here, we prepared a panel of mutants targeting residues that form the Ca2+ coordination sphere of GCPII and analyzed their structural and enzymatic properties using an array of complementary biophysical and biochemical approaches. Our data unequivocally show that even a slight disruption of the Ca2+-binding site destabilizes the three-dimensional fold of GCPII and is associated with impaired secretion, a high propensity to form nonphysiological oligomers, and an inability to bind active site-targeted ligands. Additionally, the Ca2+-binding site is critical for maintenance of the native homodimeric quaternary arrangement of GCPII, which is indispensable for its enzymatic activity. Overall, our results offer a clear picture of the importance of Ca2+ for the structural integrity and hydrolytic activity of human GCPII and by extension homologous members of the M28 zinc-dependent metallopeptidase family.

Název v anglickém jazyce

The calcium-binding site of human glutamate carboxypeptidase II is critical for dimerization, thermal stability, and enzymatic activity

Popis výsledku anglicky

Calcium ions are required for proper function of a wide spectrum of proteins within cells. X-ray crystallography of human glutamate carboxypeptidase II (GCPII) revealed the presence of a Ca2+-binding site, but its importance for the structure and function of this metallopeptidase has not been elucidated to date. Here, we prepared a panel of mutants targeting residues that form the Ca2+ coordination sphere of GCPII and analyzed their structural and enzymatic properties using an array of complementary biophysical and biochemical approaches. Our data unequivocally show that even a slight disruption of the Ca2+-binding site destabilizes the three-dimensional fold of GCPII and is associated with impaired secretion, a high propensity to form nonphysiological oligomers, and an inability to bind active site-targeted ligands. Additionally, the Ca2+-binding site is critical for maintenance of the native homodimeric quaternary arrangement of GCPII, which is indispensable for its enzymatic activity. Overall, our results offer a clear picture of the importance of Ca2+ for the structural integrity and hydrolytic activity of human GCPII and by extension homologous members of the M28 zinc-dependent metallopeptidase family.

Druh

J_imp - Článek v periodiku v databázi Web of Science

CEP obor

—

OECD FORD obor

10608 - Biochemistry and molecular biology

Projekt

Výsledek vznikl pri realizaci vícero projektů. Více informací v záložce Projekty.

Návaznosti

I - Institucionalni podpora na dlouhodoby koncepcni rozvoj vyzkumne organizace

Rok uplatnění

2018

Kód důvěrnosti údajů

S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů

Název periodika

Protein Science

ISSN

0961-8368

e-ISSN

—

Svazek periodika

27

Číslo periodika v rámci svazku

9

Stát vydavatele periodika

US - Spojené státy americké

Počet stran výsledku

10

Strana od-do

1575-1584

Kód UT WoS článku

000447779000004

EID výsledku v databázi Scopus

2-s2.0-85055040824