The Phase Transition Of Elastin-like Polypeptides With Differenttopological Structures

Elastin-like polypeptides(Elastin-like polypeptides,ELPs)are a part of artificial biopolymers that elastic andenvironmentally responsive.In this study,ELPs with various topologies weresuccessful constructed by incorporating the SpyTag/SpyCatcherand Foldonwith ELPs.The different topologicalstructures of ELPs greatly enriched the structure of traditional linear ELPs and have great significance in application.Firstly,we constructed elastin-like polypeptides(ELPs)fusion proteins with different topologicalstructures by SpyTag/SpyCatcher and investigated the effects of salts types and concentrations on the phase transition of the fusion proteins.Results showed that the phase transition temperature of ELP40 fusion protein with different typologies varied from 1to 10.9°C under different salt concentrations,which was higher than that of their corresponding linear ELPs.Amongthem,the maximum difference was observed when the concentration of Na2CO3 was 0.05 mol/L with a disparityvalue of 11.5 °C.Meanwhile,the transition temperature decreases linearly as a function of the salts concentrations.However,phase transition was observed in SpyCatcher-ELP40 only under high concentration(?0.5 mol/L)of Na2CO3 solution,and the phase transition temperature dropped dramatically to 7.3 °C.This result was obviously inconsistent withthe Hofmeister series.The possible reason was related with the amino acids with positive and negative chargesdistributed on the surface of SpyCatcher.We also addressed the possible mechanism of how this phenomenonoccurred.Secondly,we constructedthe non-covalentthree-armed star C-E-F and thethree-armed star E-F by fusing Foldon with SpyCatcher-ELP40 and ELP40 and investigated the effects of salts types and concentrations on the phase transition of the fusion proteins.We found the phase transition temperature of thethree-armed star E-F was lower than of the corresponding linear ELP40;while,the phase transition temperature of the three-armed star C-E-F increased and then decreased.At the same time,we could observe the phase transition in the three-armed star C-E-F only under the high concentration(?0.7 mol/L)of Na2CO3 solution.The phenomenon is mainly related to solvent accessible charge distribution of foldon and SpyCatcher.Finally,we constructed atadpole-like ELP40 by fusing SpyTag/SpyCatcher with ELP40.Tadpole-like ELP40 was digested with TEV protease.The TEV protease digestedproducts were assayed by SDS-PAGE and MOTIF/TOF mass spectrometry,and the result verifiesthe formation of the tadpole structure.We investigated the effects of salts types and concentrations on the phase transition of the fusion proteins.The result showed thatthe phase transition temperature of the tadpole-like ELP40 was higher than that of their corresponding linear ELP80.Weexploredthe effect of anions and cations on thephase transition temperature of the tadpole-like ELP40 in the Hofmeister series.Results showedthat the effect of anions on the phase transition temperature of tadpole-like ELP40 was consistent with the Hofmeister series,while the cation inverse the Hofmeister series.In addition,the particle size of tadpole-like ELP40 was analyzed in different salt solutions.The average particle size was between 0.6 ?m and 2 ?m,which was at the micron level and of great potentialsin application.