Since direct staining of growing hyphae with trifluoperazine (TFP) could result in the migration of protoplasm in hyphae of water mold Saprolegnia feraX, an improved staining method for fixation of growing hyphae was ...Since direct staining of growing hyphae with trifluoperazine (TFP) could result in the migration of protoplasm in hyphae of water mold Saprolegnia feraX, an improved staining method for fixation of growing hyphae was developed. When hyphae of Saprolegnia ferax were pre-fixed with paraformaldehyde, the TFP fluorescence showed a clear tip-high Ca(2+)-activated calmodulin (CaM) gradient. By indirect immunohistochemical staining, we proved that total CaM also mainly concentrated in the apex of hyphae.It is noticeble that when the hyphal tip growth ceased, the high intensity of TFP fluorescence usually disappeared in non-growing tips and became evident in the newly emerged growing tips, which implied that the concentration of CaM shifted in concord with the transfer of growing point in hyphae. 2×10(-5) mol/L CaM inhibitor TFP could reduce the tip growth rate and the percentage of growing hyphal tips to about 50%, and induce multibranching of hyphal tips. These findings strongly suggest that the high concentration of calmodulin in the hyphal tip may be involved in regulation of hyphal tip growth.展开更多
To achieve higher antibiotic streptolydigin productivity through metabolic regulation, propionate was fed during the fermentation of Streptomyces lydicus AS 4.2501. The effects of propionate feeding on streptolydigin ...To achieve higher antibiotic streptolydigin productivity through metabolic regulation, propionate was fed during the fermentation of Streptomyces lydicus AS 4.2501. The effects of propionate feeding on streptolydigin production and intracellular fluxes were investigated. The highest streptolydigin production yield of 95.10mg·L-1 was obtained when 2mmol·L-1 of sodium propionate was added at 60h of cultivation into shake-flask culture. This yield is 23.06% higher when compared to that of a batch culture without propionate supplementation. It was also found that when propionate was added, much more organic acids were excreted. Metabolic flux analysis was per-formed and it demonstrated that the carbon fluxes of the pentose phosphate pathway and the anaplerotic reaction were significantly increased after propionate feeding. The carbon flux from pyruvate to acetyl-CoA was determined to be 24.7, which was 12.27% higher than that in the batch culture. This study indicated that the glu-cose-6-phosphate and pyruvate nodes were potential bottlenecks for increasing streptolydigin productivity. Potential targets and strategies that could be manipulated through genetic and process engineering to increase the production of streptolydigin were also suggested.展开更多
文摘Since direct staining of growing hyphae with trifluoperazine (TFP) could result in the migration of protoplasm in hyphae of water mold Saprolegnia feraX, an improved staining method for fixation of growing hyphae was developed. When hyphae of Saprolegnia ferax were pre-fixed with paraformaldehyde, the TFP fluorescence showed a clear tip-high Ca(2+)-activated calmodulin (CaM) gradient. By indirect immunohistochemical staining, we proved that total CaM also mainly concentrated in the apex of hyphae.It is noticeble that when the hyphal tip growth ceased, the high intensity of TFP fluorescence usually disappeared in non-growing tips and became evident in the newly emerged growing tips, which implied that the concentration of CaM shifted in concord with the transfer of growing point in hyphae. 2×10(-5) mol/L CaM inhibitor TFP could reduce the tip growth rate and the percentage of growing hyphal tips to about 50%, and induce multibranching of hyphal tips. These findings strongly suggest that the high concentration of calmodulin in the hyphal tip may be involved in regulation of hyphal tip growth.
基金Supported by the National Natural Science Foundation of China (No.20425620)the Program for Changjiang Scholars andInnovative Research Team in University from the Ministry of Education of China.
文摘To achieve higher antibiotic streptolydigin productivity through metabolic regulation, propionate was fed during the fermentation of Streptomyces lydicus AS 4.2501. The effects of propionate feeding on streptolydigin production and intracellular fluxes were investigated. The highest streptolydigin production yield of 95.10mg·L-1 was obtained when 2mmol·L-1 of sodium propionate was added at 60h of cultivation into shake-flask culture. This yield is 23.06% higher when compared to that of a batch culture without propionate supplementation. It was also found that when propionate was added, much more organic acids were excreted. Metabolic flux analysis was per-formed and it demonstrated that the carbon fluxes of the pentose phosphate pathway and the anaplerotic reaction were significantly increased after propionate feeding. The carbon flux from pyruvate to acetyl-CoA was determined to be 24.7, which was 12.27% higher than that in the batch culture. This study indicated that the glu-cose-6-phosphate and pyruvate nodes were potential bottlenecks for increasing streptolydigin productivity. Potential targets and strategies that could be manipulated through genetic and process engineering to increase the production of streptolydigin were also suggested.
