Chemical proteomics study of the anti-metastasis activity of andrographolide on B16 melanoma cells
碩士 === 國立陽明大學 === 生物藥學研究所 === 100 === Serine hydrolase family, estimated as 1% of human genome, is one of the largest and diverse enzyme classes in mammalian proteome, which participates in essential physiological functions. Besides, the activity levels of several serine hydrolases have been recentl...
Main Authors: | , |
---|---|
Other Authors: | |
Format: | Others |
Language: | zh-TW |
Published: |
2012
|
Online Access: | http://ndltd.ncl.edu.tw/handle/33991252848652125984 |
id |
ndltd-TW-100YM005603006 |
---|---|
record_format |
oai_dc |
spelling |
ndltd-TW-100YM0056030062015-10-13T21:22:39Z http://ndltd.ncl.edu.tw/handle/33991252848652125984 Chemical proteomics study of the anti-metastasis activity of andrographolide on B16 melanoma cells 穿心蓮內酯抑制小鼠黑色素瘤細胞轉移能力 之化學蛋白體學研究 Ting-Yun Huang 黃庭筠 碩士 國立陽明大學 生物藥學研究所 100 Serine hydrolase family, estimated as 1% of human genome, is one of the largest and diverse enzyme classes in mammalian proteome, which participates in essential physiological functions. Besides, the activity levels of several serine hydrolases have been recently reported to correlate with tumor progression or migration. However, more than half members of this enzyme family remain unclear for their substrates and physiological function. Therefore, systematic identification of disease-related serine hydrolases is very important. Andrographolide, a major chemical ingredient of Chinese herb Andrographis panicμlata, was previously shown to exert its anticancer activity through cell growth inhibition, apoptosis induction, antiangiogenesis and metastasis inhibition. In previous mechanistic studies, andrographolide suppressed cell invasion of colorectal cancer cells (CT26 and HT29) via inhibiting MMP2 activity and ERK expression. However, whether the activity of serine hydrolase was involved in andrographolide-mediated inhibition of cancer cell migration has not been reported. Therefore, my thesis study intended to investigate whether serine hydrolases participate in the anti-migration activity of andrographolide. A chemical proteomics approach was applied to analyze serine hydrolases and to identify alteration of activity profiles of serine hydrolases in two mouse melanoma cells, B16F1 and B16F10 which differ in migration ability, upon andrographolide treatment. The biotin-tagged probes further used to isolate serine hydrolases whose activities were affected by andrographolide, which woμld increase our understanding towards the mechanism and involved pathways of andrographolide-induced suppression of cancer cell migration. Through collaboration with Professor Chung-Ming Sun from Dept. of Applied Chemistry in NCTU and Professor Lee-Chiang Lo from Dept. of Chemistry in NTU, two types of serine hydrolase probes carrying either biotin or fluorophore tags, namely FP-edo-biotin, FP-edo-fluorescein and FP-benzyl-biotin, FP-benzyl-BODIPY, were prepared. I first used the poly-3-hydroxybutyrate depolymerase (PhaZ) from Bacillus thuringiensis to examine the probe efficiency at diverse reaction concentration and time. Moreover, the fluorescent probes were used to monitor alteration of the fluorescence-based activity profiles of serine hydrolases in melanoma cells, B16F1 and B16F10. A fluorescence band was observed to be correlated with difference of cell migration between melanoma cells. Interestingly, this signal was suppressed upon andrographolide-treatment. Finally, analyses of mass spectrometry and Western blot demonstrated that such fluorescence change was likely resμlted from activity alteration of monoacylglycerol lipase (MAGL). In summary, my study applied the chemical proteomics to discover that andrographolide coμld inhibit the activity of MAGL in melanoma cell, providing a putative mechanism for the anti-migration activity of andrographolide. Chao-Hsiung Lin 林照雄 2012 學位論文 ; thesis 59 zh-TW |
collection |
NDLTD |
language |
zh-TW |
format |
Others
|
sources |
NDLTD |
description |
碩士 === 國立陽明大學 === 生物藥學研究所 === 100 === Serine hydrolase family, estimated as 1% of human genome, is one of the largest and diverse enzyme classes in mammalian proteome, which participates in essential physiological functions. Besides, the activity levels of several serine hydrolases have been recently reported to correlate with tumor progression or migration. However, more than half members of this enzyme family remain unclear for their substrates and physiological function. Therefore, systematic identification of disease-related serine hydrolases is very important. Andrographolide, a major chemical ingredient of Chinese herb Andrographis panicμlata, was previously shown to exert its anticancer activity through cell growth inhibition, apoptosis induction, antiangiogenesis and metastasis inhibition. In previous mechanistic studies, andrographolide suppressed cell invasion of colorectal cancer cells (CT26 and HT29) via inhibiting MMP2 activity and ERK expression. However, whether the activity of serine hydrolase was involved in andrographolide-mediated inhibition of cancer cell migration has not been reported. Therefore, my thesis study intended to investigate whether serine hydrolases participate in the anti-migration activity of andrographolide. A chemical proteomics approach was applied to analyze serine hydrolases and to identify alteration of activity profiles of serine hydrolases in two mouse melanoma cells, B16F1 and B16F10 which differ in migration ability, upon andrographolide treatment. The biotin-tagged probes further used to isolate serine hydrolases whose activities were affected by andrographolide, which woμld increase our understanding towards the mechanism and involved pathways of andrographolide-induced suppression of cancer cell migration. Through collaboration with Professor Chung-Ming Sun from Dept. of Applied Chemistry in NCTU and Professor Lee-Chiang Lo from Dept. of Chemistry in NTU, two types of serine hydrolase probes carrying either biotin or fluorophore tags, namely FP-edo-biotin, FP-edo-fluorescein and FP-benzyl-biotin, FP-benzyl-BODIPY, were prepared. I first used the poly-3-hydroxybutyrate depolymerase (PhaZ) from Bacillus thuringiensis to examine the probe efficiency at diverse reaction concentration and time. Moreover, the fluorescent probes were used to monitor alteration of the fluorescence-based activity profiles of serine hydrolases in melanoma cells, B16F1 and B16F10. A fluorescence band was observed to be correlated with difference of cell migration between melanoma cells. Interestingly, this signal was suppressed upon andrographolide-treatment. Finally, analyses of mass spectrometry and Western blot demonstrated that such fluorescence change was likely resμlted from activity alteration of monoacylglycerol lipase (MAGL). In summary, my study applied the chemical proteomics to discover that andrographolide coμld inhibit the activity of MAGL in melanoma cell, providing a putative mechanism for the anti-migration activity of andrographolide.
|
author2 |
Chao-Hsiung Lin |
author_facet |
Chao-Hsiung Lin Ting-Yun Huang 黃庭筠 |
author |
Ting-Yun Huang 黃庭筠 |
spellingShingle |
Ting-Yun Huang 黃庭筠 Chemical proteomics study of the anti-metastasis activity of andrographolide on B16 melanoma cells |
author_sort |
Ting-Yun Huang |
title |
Chemical proteomics study of the anti-metastasis activity of andrographolide on B16 melanoma cells |
title_short |
Chemical proteomics study of the anti-metastasis activity of andrographolide on B16 melanoma cells |
title_full |
Chemical proteomics study of the anti-metastasis activity of andrographolide on B16 melanoma cells |
title_fullStr |
Chemical proteomics study of the anti-metastasis activity of andrographolide on B16 melanoma cells |
title_full_unstemmed |
Chemical proteomics study of the anti-metastasis activity of andrographolide on B16 melanoma cells |
title_sort |
chemical proteomics study of the anti-metastasis activity of andrographolide on b16 melanoma cells |
publishDate |
2012 |
url |
http://ndltd.ncl.edu.tw/handle/33991252848652125984 |
work_keys_str_mv |
AT tingyunhuang chemicalproteomicsstudyoftheantimetastasisactivityofandrographolideonb16melanomacells AT huángtíngyún chemicalproteomicsstudyoftheantimetastasisactivityofandrographolideonb16melanomacells AT tingyunhuang chuānxīnliánnèizhǐyìzhìxiǎoshǔhēisèsùliúxìbāozhuǎnyínénglìzhīhuàxuédànbáitǐxuéyánjiū AT huángtíngyún chuānxīnliánnèizhǐyìzhìxiǎoshǔhēisèsùliúxìbāozhuǎnyínénglìzhīhuàxuédànbáitǐxuéyánjiū |
_version_ |
1718062169458737152 |