Please use this identifier to cite or link to this item: http://hdl.handle.net/11455/38077
標題: A continuous spectrophotometric assay method for peptidylarginine deiminase type 4 activity
作者: Liao, Y.F.
洪慧芝
Hsieh, H.C.
Liu, G.Y.
Hung, H.C.
關鍵字: peptidylarginine deiminase
glutamate-dehydrogenase
spectrophotometric
continuous assay
gene organization
expression analysis
arginine deiminase
molecular-cloning
cdna cloning
human skin
performance
hydrolysis
citrulline
enzymes
期刊/報告no:: Analytical Biochemistry, Volume 347, Issue 2, Page(s) 176-181.
摘要: A simple, continuous spectrophotometric assay for peptidylarginine deiminase (PAD) is described. Deimination of peptidylarginine results in the formation of peptidylcitrulline and ammonia. The ammonia released during peptidylarginine hydrolysis is coupled to the glutamate-dehydrogenase-catalyzed reductive amination of of alpha-ketoglutarate to glutamate and reduced nicotinamide adenine dinucleotide (NADH) oxidation. The disappearance of absorbance at 340 nm due to NADH oxidation is continuously measured. The specific activity obtained by this new protocol for highly purified human PAD is comparable to that obtained by a commonly used colorimetric procedure, which measures the ureido group of peptidylcitrulline by coupling with diacetyl monoxime. The present continuous spectrophotometric method is highly sensitive and accurate and is thus suitable for enzyme kinetic analysis of PAD. The Ca2+ concentration for half-maximal activity of PAD obtained by this method is comparable to that previously obtained by the colorimetric procedure. (c) 2005 Elsevier Inc. All rights reserved.
URI: http://hdl.handle.net/11455/38077
ISSN: 0003-2697
文章連結: http://dx.doi.org/10.1016/j.ab.2005.09.027
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