Conserved lysine 79 is important for activity of ecto-nucleoside triphosphate diphosphohydrolase 3 (NTPDase3)

Cell membrane-bound ecto-nucleoside triphosphate diphosphohydrolases (NTPDases) are homooligomeric, with native quaternary structure required for maximal enzyme activity. In this study, we mutated lysine 79 in human ecto-nucleoside triphosphate diphosphohydrolase 3 (NTPDase3). The residue correspond...

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Main Authors:	Basu, Saswata, Murphy-Piedmonte, Deirdre M., Kirley, Terence L.
Format:	Online
Language:	English
Published:	Springer Netherlands 2004
Online Access:	https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2096571/

id	pubmed-2096571
recordtype	oai_dc
spelling	pubmed-20965712008-02-27 Conserved lysine 79 is important for activity of ecto-nucleoside triphosphate diphosphohydrolase 3 (NTPDase3) Basu, Saswata Murphy-Piedmonte, Deirdre M. Kirley, Terence L. Original Paper Cell membrane-bound ecto-nucleoside triphosphate diphosphohydrolases (NTPDases) are homooligomeric, with native quaternary structure required for maximal enzyme activity. In this study, we mutated lysine 79 in human ecto-nucleoside triphosphate diphosphohydrolase 3 (NTPDase3). The residue corresponding to lysine 79 in NTPDase3 is conserved in all known cell surface membrane NTPDases (NTPDase1, 2, 3, and 8), but not in the soluble, monomeric NTPDases (NTPDase5 and 6), or in the intracellular, two transmembrane NTPDases (NTPDase4 and 7). This conserved lysine is located between apyrase conserved region 1 (ACR1) and an invariant glycosylation site (N81), in a region previously hypothesized to be important for NTPDase3 oligomeric structure. This lysine residue was mutated to several different amino acids, and all mutants displayed substantially decreased nucleotidase activities. A basic amino acid at this position was found to be important for the increase of nucleotidase activity observed after treatment with the lectin, concanavalin A. After solubilization with Triton X-100, mutants showed little or no decrease in activity, unlike the wild-type enzyme, suggesting that the lysine at this position may be important for maintaining proper folding and for stabilizing the quaternary structure. However, mutation at this site did not result in global changes in tertiary or quaternary structure as measured by Cibacron blue binding, chemical cross linking, and native gel electrophoretic analysis, leaving open the possibility of other mechanisms by which mutation of this conserved lysine residue might decrease enzyme activity. Springer Netherlands 2004-12 /pmc/articles/PMC2096571/ /pubmed/18404400 http://dx.doi.org/10.1007/s11302-004-4741-8 Text en © Springer 2004
repository_type	Open Access Journal
institution_category	Foreign Institution
institution	US National Center for Biotechnology Information
building	NCBI PubMed
collection	Online Access
language	English
format	Online
author	Basu, Saswata Murphy-Piedmonte, Deirdre M. Kirley, Terence L.
spellingShingle	Basu, Saswata Murphy-Piedmonte, Deirdre M. Kirley, Terence L. Conserved lysine 79 is important for activity of ecto-nucleoside triphosphate diphosphohydrolase 3 (NTPDase3)
author_facet	Basu, Saswata Murphy-Piedmonte, Deirdre M. Kirley, Terence L.
author_sort	Basu, Saswata
title	Conserved lysine 79 is important for activity of ecto-nucleoside triphosphate diphosphohydrolase 3 (NTPDase3)
title_short	Conserved lysine 79 is important for activity of ecto-nucleoside triphosphate diphosphohydrolase 3 (NTPDase3)
title_full	Conserved lysine 79 is important for activity of ecto-nucleoside triphosphate diphosphohydrolase 3 (NTPDase3)
title_fullStr	Conserved lysine 79 is important for activity of ecto-nucleoside triphosphate diphosphohydrolase 3 (NTPDase3)
title_full_unstemmed	Conserved lysine 79 is important for activity of ecto-nucleoside triphosphate diphosphohydrolase 3 (NTPDase3)
title_sort	conserved lysine 79 is important for activity of ecto-nucleoside triphosphate diphosphohydrolase 3 (ntpdase3)
description	Cell membrane-bound ecto-nucleoside triphosphate diphosphohydrolases (NTPDases) are homooligomeric, with native quaternary structure required for maximal enzyme activity. In this study, we mutated lysine 79 in human ecto-nucleoside triphosphate diphosphohydrolase 3 (NTPDase3). The residue corresponding to lysine 79 in NTPDase3 is conserved in all known cell surface membrane NTPDases (NTPDase1, 2, 3, and 8), but not in the soluble, monomeric NTPDases (NTPDase5 and 6), or in the intracellular, two transmembrane NTPDases (NTPDase4 and 7). This conserved lysine is located between apyrase conserved region 1 (ACR1) and an invariant glycosylation site (N81), in a region previously hypothesized to be important for NTPDase3 oligomeric structure. This lysine residue was mutated to several different amino acids, and all mutants displayed substantially decreased nucleotidase activities. A basic amino acid at this position was found to be important for the increase of nucleotidase activity observed after treatment with the lectin, concanavalin A. After solubilization with Triton X-100, mutants showed little or no decrease in activity, unlike the wild-type enzyme, suggesting that the lysine at this position may be important for maintaining proper folding and for stabilizing the quaternary structure. However, mutation at this site did not result in global changes in tertiary or quaternary structure as measured by Cibacron blue binding, chemical cross linking, and native gel electrophoretic analysis, leaving open the possibility of other mechanisms by which mutation of this conserved lysine residue might decrease enzyme activity.
publisher	Springer Netherlands
publishDate	2004
url	https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2096571/
_version_	1611408394294394880

Conserved lysine 79 is important for activity of ecto-nucleoside triphosphate diphosphohydrolase 3 (NTPDase3)

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