AIM: To investigate the hepatoprotective effects of phycocyanobilin(PCB) in reducing hepatic injury and accelerating hepatocyte proliferation following carbon tetrachloride(CCl4) treatment.METHODS: C57BL/6 mice were o...AIM: To investigate the hepatoprotective effects of phycocyanobilin(PCB) in reducing hepatic injury and accelerating hepatocyte proliferation following carbon tetrachloride(CCl4) treatment.METHODS: C57BL/6 mice were orally administered PCB 100 mg/kg for 4 d after CCl4 injection, and then the serum and liver tissue of the mice were collected at days 1, 2, 3, 5 and 7 after CCl4 treatment. A series of evaluations were performed to identify the curative effects on liver injury and recovery. Aspartate aminotransferase(AST), alanine aminotransferase(ALT), albumin and superoxide dismutase(SOD) were detected to indirectly assess the anti-inflammatory effects of PCB. Meanwhile, we detected the expressions of hepatocyte growth factor, transforming growth factor alpha(TGF-α), TGF-β, tumor necrosis factor-alpha(TNF-α) and interleukin-6(IL-6), the factors which are associated with inflammation and liver regeneration. The protein expressions of proliferating cell nuclear antigen(PCNA), TNF-α and cytochrome C were detected by western blot. Furthermore, the survivalrates were analyzed of mice which were administered a lethal dose of CCl4(2.6 mg/kg)with or without PCB.RESULTS:In our research,PCB showed a strongly anti-inflammatory effect on CCl4-induced liver injury in mice.The ALT was significantly decreased after CCl4 treatment from day 1(P<0.01)and the AST was significantly decreased from day 2(P<0.001).Both albumin and liver SOD were increased from day2(P<0.001 and P<0.01),but serum SOD levels did not show a significant increase(P>0.05).PCB protected the structure of liver from the injury by CCl4.TUNEL assay showed that PCB dramatically reduced the number of apoptotic cells after CCl4 treatment compared to the control(101.0±25.4 vs 25.7±6.4,P<0.01).The result of western blotting showed that PCB could increase PCNA expression,decrease TNF-αand cytochrome C expression.Furthermore,data shows that PCB could improve the survival rate of acute liver failure(ALF)mice which were injected with a lethal dose of CCl4(60.0%vs 20.0%).CONCLUSION:Our study indicated that PCB could be an ideal candidate for reversing acute liver injury or ALF.展开更多
The recombinant expression of phycocyanobilin(PCB)was carried out in Escherichia coli,and the best fermentation conditions of recombinant E.coli biosynthesized PCB are optimized in the response surface methodology to ...The recombinant expression of phycocyanobilin(PCB)was carried out in Escherichia coli,and the best fermentation conditions of recombinant E.coli biosynthesized PCB are optimized in the response surface methodology to improve PCB production.The recombinant PCB is extracted,isolated,and purified by methanol and chloroform extraction.Recombinant PCB is validated in UV-vis spectroscopy,high-pressure liquid chromatography,and mass spectrometry.In addition,the anti-oxidant activities of the recombinant PCB are determined.The best induction conditions that optimized by Design Expert 8.0 software include:lactose concentration 4 mmol/L,induction temperature 24.69℃,induction time 4.6 h,and induction duration 13.57 h,under which the PCB expression level reached approximately 13 mg PCB/L,which is more than four times of previously reported 3 mg PCB/L.The maximum absorption peak of the recombinant PCB is located at 680 nm with a high fluorescence intensity of 470 nm.The recombinant PCB has a good ability to scavenge 2,2-diphenyl-1-picrylhydrazyl(DPPH)free radicals.展开更多
Phycocyanobilin(PCB)is widely used in healthcare,food processing,and cosmetics.Escherichia coli is the common engineered bacterium used to produce PCB.However,it still suffers from low production level,pre-cursor defi...Phycocyanobilin(PCB)is widely used in healthcare,food processing,and cosmetics.Escherichia coli is the common engineered bacterium used to produce PCB.However,it still suffers from low production level,pre-cursor deficiency,and low catalytic efficiency.In this study,a highly efficient PCB-producing strain was created.First,chassis strains and enzyme sources were screened,and copy numbers were optimized,affording a PCB titer of 9.1 mg/L.Most importantly,the rate-limiting steps of the PCB biosynthetic pathway were determined,and the supply of precursors necessary for PCB synthesis was increased from endogenous sources,affording a titer of 21.4 mg/L.Then,the key enzymes for PCB synthesis,HO1 and PcyA,were assembled into a multi-enzyme complex using the short peptide tag RIAD-RIDD,and 23.5 mg/L of PCB was obtained.Finally,the basic con-ditions for PCB fermentation were initially determined in 250 mL shake flasks and a 5-L bioreactor to obtain higher titers of PCB.The final titer of PCB reached 147.0 mg/L,which is the highest reported titer of PCB so far.This research provided the foundation for the industrial production of PCB and its derivatives.展开更多
Phycobiliproteins, part of the photosynthetic apparatus in algae, are a group of important absorbers of light, in which the functional groups for light absorption are linear tetrapyrrole chromophores. In order to inte...Phycobiliproteins, part of the photosynthetic apparatus in algae, are a group of important absorbers of light, in which the functional groups for light absorption are linear tetrapyrrole chromophores. In order to interpret the photobiological phenomenon of the phycobiliproteins, it is important to investigate the chemical properties and the formation of the derivatives of the chromophores.展开更多
Arthrospira platensis is an economically important cyanobacterium; and it has been used widely in food and pharmaceutical industries. The phycocyanin(PC) from A. platensis is extremely valuable in medicine and molecul...Arthrospira platensis is an economically important cyanobacterium; and it has been used widely in food and pharmaceutical industries. The phycocyanin(PC) from A. platensis is extremely valuable in medicine and molecular biology due to its antioxidation and anti-tumoring activity and applicability as fluorescence protein tag. In present study, two recombinant plasmids, one contained the phycocyanobilin(PCB)-producing genes(hox1 and pcyA) while the other contained the phycobiliprotein gene(cpcB) and the lyase gene(either cpcS/U or cpcT), were constructed and synchronically transferred into E. coli in order to test the the activities of relevant lyases for catalysing PCB addition to CpcB during synthesizing fluorescent PC holo-β-subunit(β-PC) of A. platensis. As was evidenced by the fluorescence emitted at a peak specific for PC, CpcB was successfully synthesized in E. coli, to which co-expressed PCBs attached though at a relatively low efficiency. The results showed that the attachment of PCBs to CpcB were carried out mainly by co-expressed CpcS/U but CpcB also showed some autocatalytic activity. Currently, no CpcT activity was detected in this E. coli expression system. Further studies will be conducted to improve the efficiency of fluorescent PC synthesis in E. coli.展开更多
A spectral method to investigate the effect of Fe3+, Fe2+ on the thermostability ofphycocyanin (PC) of Spirulina maxima showed that iron ions prevent decrease of visible light absorbanceand fluorescence intensity of P...A spectral method to investigate the effect of Fe3+, Fe2+ on the thermostability ofphycocyanin (PC) of Spirulina maxima showed that iron ions prevent decrease of visible light absorbanceand fluorescence intensity of PC. Increase in denaturation temperature caused by Fe3+ was observed bythe micro - differential scanning calorimetric method. All results showed iron ions maintain the aggrega-tion stability of the PC. The absorption spectrum of phycocyanobilin (PCB, a prosthetic group of PC) withFe3+ in chloroform was quite different from that of free PCB.展开更多
基金Supported by Zhanjiang Key Laboratory of Hepatobiliary Diseases,No.2013A402-4The Medical Research Funding of Affiliated Hospital of Guangdong Medical College,No.QK1319The Medical Research Funding of Guangdong Province,No.B2014306,China
文摘AIM: To investigate the hepatoprotective effects of phycocyanobilin(PCB) in reducing hepatic injury and accelerating hepatocyte proliferation following carbon tetrachloride(CCl4) treatment.METHODS: C57BL/6 mice were orally administered PCB 100 mg/kg for 4 d after CCl4 injection, and then the serum and liver tissue of the mice were collected at days 1, 2, 3, 5 and 7 after CCl4 treatment. A series of evaluations were performed to identify the curative effects on liver injury and recovery. Aspartate aminotransferase(AST), alanine aminotransferase(ALT), albumin and superoxide dismutase(SOD) were detected to indirectly assess the anti-inflammatory effects of PCB. Meanwhile, we detected the expressions of hepatocyte growth factor, transforming growth factor alpha(TGF-α), TGF-β, tumor necrosis factor-alpha(TNF-α) and interleukin-6(IL-6), the factors which are associated with inflammation and liver regeneration. The protein expressions of proliferating cell nuclear antigen(PCNA), TNF-α and cytochrome C were detected by western blot. Furthermore, the survivalrates were analyzed of mice which were administered a lethal dose of CCl4(2.6 mg/kg)with or without PCB.RESULTS:In our research,PCB showed a strongly anti-inflammatory effect on CCl4-induced liver injury in mice.The ALT was significantly decreased after CCl4 treatment from day 1(P<0.01)and the AST was significantly decreased from day 2(P<0.001).Both albumin and liver SOD were increased from day2(P<0.001 and P<0.01),but serum SOD levels did not show a significant increase(P>0.05).PCB protected the structure of liver from the injury by CCl4.TUNEL assay showed that PCB dramatically reduced the number of apoptotic cells after CCl4 treatment compared to the control(101.0±25.4 vs 25.7±6.4,P<0.01).The result of western blotting showed that PCB could increase PCNA expression,decrease TNF-αand cytochrome C expression.Furthermore,data shows that PCB could improve the survival rate of acute liver failure(ALF)mice which were injected with a lethal dose of CCl4(60.0%vs 20.0%).CONCLUSION:Our study indicated that PCB could be an ideal candidate for reversing acute liver injury or ALF.
基金Supported by the National Natural Science Foundation of China(No.41906109)the Key Research and Development Program of Yantai(No.Y933021011)
文摘The recombinant expression of phycocyanobilin(PCB)was carried out in Escherichia coli,and the best fermentation conditions of recombinant E.coli biosynthesized PCB are optimized in the response surface methodology to improve PCB production.The recombinant PCB is extracted,isolated,and purified by methanol and chloroform extraction.Recombinant PCB is validated in UV-vis spectroscopy,high-pressure liquid chromatography,and mass spectrometry.In addition,the anti-oxidant activities of the recombinant PCB are determined.The best induction conditions that optimized by Design Expert 8.0 software include:lactose concentration 4 mmol/L,induction temperature 24.69℃,induction time 4.6 h,and induction duration 13.57 h,under which the PCB expression level reached approximately 13 mg PCB/L,which is more than four times of previously reported 3 mg PCB/L.The maximum absorption peak of the recombinant PCB is located at 680 nm with a high fluorescence intensity of 470 nm.The recombinant PCB has a good ability to scavenge 2,2-diphenyl-1-picrylhydrazyl(DPPH)free radicals.
基金supported by the National Key Research and Development Program of China(2019YFA0904800)the Foundation for Innovative Research Groups of the National Natural Science Foundation of China(32021005).
文摘Phycocyanobilin(PCB)is widely used in healthcare,food processing,and cosmetics.Escherichia coli is the common engineered bacterium used to produce PCB.However,it still suffers from low production level,pre-cursor deficiency,and low catalytic efficiency.In this study,a highly efficient PCB-producing strain was created.First,chassis strains and enzyme sources were screened,and copy numbers were optimized,affording a PCB titer of 9.1 mg/L.Most importantly,the rate-limiting steps of the PCB biosynthetic pathway were determined,and the supply of precursors necessary for PCB synthesis was increased from endogenous sources,affording a titer of 21.4 mg/L.Then,the key enzymes for PCB synthesis,HO1 and PcyA,were assembled into a multi-enzyme complex using the short peptide tag RIAD-RIDD,and 23.5 mg/L of PCB was obtained.Finally,the basic con-ditions for PCB fermentation were initially determined in 250 mL shake flasks and a 5-L bioreactor to obtain higher titers of PCB.The final titer of PCB reached 147.0 mg/L,which is the highest reported titer of PCB so far.This research provided the foundation for the industrial production of PCB and its derivatives.
基金Project supported by the National Natural Science Foundation of China and the United Analytical Testing Center in Zhongguancun Area.
文摘Phycobiliproteins, part of the photosynthetic apparatus in algae, are a group of important absorbers of light, in which the functional groups for light absorption are linear tetrapyrrole chromophores. In order to interpret the photobiological phenomenon of the phycobiliproteins, it is important to investigate the chemical properties and the formation of the derivatives of the chromophores.
基金supported by the National Science and Technology Major Project of China (2008ZX08001-004)
文摘Arthrospira platensis is an economically important cyanobacterium; and it has been used widely in food and pharmaceutical industries. The phycocyanin(PC) from A. platensis is extremely valuable in medicine and molecular biology due to its antioxidation and anti-tumoring activity and applicability as fluorescence protein tag. In present study, two recombinant plasmids, one contained the phycocyanobilin(PCB)-producing genes(hox1 and pcyA) while the other contained the phycobiliprotein gene(cpcB) and the lyase gene(either cpcS/U or cpcT), were constructed and synchronically transferred into E. coli in order to test the the activities of relevant lyases for catalysing PCB addition to CpcB during synthesizing fluorescent PC holo-β-subunit(β-PC) of A. platensis. As was evidenced by the fluorescence emitted at a peak specific for PC, CpcB was successfully synthesized in E. coli, to which co-expressed PCBs attached though at a relatively low efficiency. The results showed that the attachment of PCBs to CpcB were carried out mainly by co-expressed CpcS/U but CpcB also showed some autocatalytic activity. Currently, no CpcT activity was detected in this E. coli expression system. Further studies will be conducted to improve the efficiency of fluorescent PC synthesis in E. coli.
文摘A spectral method to investigate the effect of Fe3+, Fe2+ on the thermostability ofphycocyanin (PC) of Spirulina maxima showed that iron ions prevent decrease of visible light absorbanceand fluorescence intensity of PC. Increase in denaturation temperature caused by Fe3+ was observed bythe micro - differential scanning calorimetric method. All results showed iron ions maintain the aggrega-tion stability of the PC. The absorption spectrum of phycocyanobilin (PCB, a prosthetic group of PC) withFe3+ in chloroform was quite different from that of free PCB.
