使用4-磺基苯基异硫氰酸酯标记和质谱法确定链球菌溶解肽聚糖水解酶溶链菌酶A的作用位点。
Use of 4-sulfophenyl isothiocyanate labeling and mass spectrometry to determine the site of action of the streptococcolytic peptidoglycan hydrolase zoocin A.
作者信息
Gargis Shaw R, Heath Harry E, Heath Lucie S, Leblanc Paul A, Simmonds Robin S, Abbott Brian D, Timkovich Russell, Sloan Gary L
机构信息
Department of Biological Sciences, Box 870334, The University of Alabama, Tuscaloosa, AL 35487-0334, USA.
出版信息
Appl Environ Microbiol. 2009 Jan;75(1):72-7. doi: 10.1128/AEM.01647-08. Epub 2008 Oct 31.
Abstract
Zoocin A is a streptococcolytic peptidoglycan hydrolase with an unknown site of action that is produced by Streptococcus equi subsp. zooepidemicus 4881. Zoocin A has now been determined to be a d-alanyl-l-alanine endopeptidase by digesting susceptible peptidoglycan with a combination of mutanolysin and zoocin A, separating the resulting muropeptides by reverse-phase high-pressure liquid chromatography, and analyzing them by mass spectrometry (MS) in both the positive- and negative-ion modes to determine their compositions. In order to distinguish among possible structures for these muropeptides, they were N-terminally labeled with 4-sulfophenyl isothiocyanate (SPITC) and analyzed by tandem MS in the negative-ion mode. This novel application of SPITC labeling and MS/MS analysis can be used to analyze the structure of peptidoglycans and to determine the sites of action of other peptidoglycan hydrolases.
摘要
马链菌素A是一种具有未知作用位点的链球菌溶菌肽聚糖水解酶，由马链球菌兽疫亚种4881产生。通过用变溶菌素和马链菌素A组合消化敏感肽聚糖、利用反相高压液相色谱分离产生的胞壁肽并通过正离子和负离子模式的质谱（MS）分析它们的组成，现已确定马链菌素A是一种D-丙氨酰-L-丙氨酸内肽酶。为了区分这些胞壁肽的可能结构，它们用4-磺基苯基异硫氰酸酯（SPITC）进行N端标记，并通过负离子模式的串联质谱进行分析。SPITC标记和MS/MS分析的这种新应用可用于分析肽聚糖的结构并确定其他肽聚糖水解酶的作用位点。
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