Extracellular Phosphoprotein Regulation is Affected by Culture System Scale-Down.
Biochimica et biophysica acta G, General subjects/Biochimica et biophysica acta General subjects (Online)(2022)
摘要
BACKGROUND:Phosphorylated proteins are known to be present in multiple body fluids in normal conditions, and abnormally accumulated under some pathological conditions. The biological significance of their role in the extracellular space has started being elucidated only recently, for example in bone mineralization, neural development, and coagulation. Here, we address some criticalities of conventional culture systems for the study of the extracellular regulation of phosphorylation.METHODS:We make use of microfluidics to scale-down the culture volume to a size comparable to the interstitial spaces occurring in vivo. The phosphoprotein content of conditioned media was analyzed by a colorimetric assay that detects global phosphorylation.RESULTS:We found that miniaturization of the culture system increases phosphoprotein accumulation. Moreover, we demonstrated that in conventional culture systems dilution affects the extent of the phosphorylation reactions occurring within the extracellular space. On the other hand, in microfluidics the phosphorylation status was not affected by addition of adenosine triphosphate (ATP) and FAM20C Golgi Associated Secretory Pathway Kinase (FAM20C) ectokinase, as if their concentration was already not limiting for the phosphorylation reaction to occur.CONCLUSIONS:The volume of the extracellular environment plays a role in the process of extracellular phosphorylation due to its effect on the concentration of substrates, enzymes and co-factors.GENERAL SIGNIFICANCE:Thus, the biological role of extracellular phosphoregulation may be better appreciated within a microfluidic culture system.
更多查看译文
关键词
Phosphorylation,Extracellular,Microfluidics,Ectokinase,Ectophosphatase,Phosphoprotein
AI 理解论文
溯源树
样例
生成溯源树,研究论文发展脉络
Chat Paper
正在生成论文摘要