Insight into Substrate Preference of Two Chimeric Esterases by Combining Experiment and Molecular Simulation

Better understanding of the relationship between the substrate preference and structural module of esterases is helpful to novel enzyme development. For this purpose, two chimeric esterases AAM7 and PAR, constructed via domain swapping between two ancient thermophilic esterases, were investigated on their molecular simulation（including homology modeling, substrates docking and substrate binding affinity validation） and enzymatic assay（specific activities and activation energies calculating）. Our results indicate that the factors contributing to the substrate preference of many enzymes especially the broad-specificity enzymes like esterases are multiple and complicated, the substrate binding domains or binding pockets are important but not the only factor for substrate preference.

Effects of Sand Grain Size on Habitat Selection in Steppe Toad-headed Lizard(Phrynocephalus frontalis)

To understand the effects of sand grain size on habitat selection, we conducted a field study on the steppe toad-headed lizard in Hunshandake Desert northern China. Methods of mark-recapture and transect survey were used to investigate the density of steppe toad-headed lizards and the environmental variables. The comparison on lizard densities among the habitats with different environmental factors revealed that： 1） population density of the steppe toad-headed lizard differed significantly among the habitats with different sand grain size indexes（SGSIs, representing roughness of sand substrate）： the highest lizard density was found in the group with an SGSI of 〉 0.30, whereas the lowest density was found in the group with an SGSI of 0–0.15; and 2） vegetation cover, soil moisture, invertebrate diversity index, and abundance had no significant effects on the lizard density. These results implied that the sand grain size was the most important determinant of habitat selection for steppe toad-headed lizards in Hunshandake Desert. Steppe toad-headed lizards could avoid structural habitats that have negative effects on their maximal sprinting capabilities. Considering the changing sand grain size in the development phase of sand dunes, the sand lizard could be used as an indicator of the process of desertification.

Histone methyltransferase TXRl is required for both H3 and H3.3lysine 27 methylation in the well-known ciliated protist Tetrahymena thermophila

DNA replication elongation is tightly controlled by histone-modifying enzymes.Our previous studies showed that the histone methytransferase TXRl(Tetrahymena Trithorax related protein 1) specifically catalyzes H3K27 monomethylation and affects DNA replication elongation in Tetrahymena thermophila.In this study,we investigated whether TXRl has a substrate preference to the canonical H3 over the replacement variant H3.3.We demonstrated by histone mutagenesis that K27 Q mutation in H3.3further aggravated the replication stress phenotype of K27 Q mutation in canonical H3,supporting H3.3 as a physiologically relevant substrate of TXRl.This result is in apparent contrast to the strong preference for canonical H3 recently reported in Arabidopsis homologues ATXR5 and ATXR6,and further corroborates the role of TXRl in DNA replication.

Purification and studies on characteristics of cholinesterases from Daphnia magna

Due to their significant value in both economy and ecology,Daphnia had long been employed to investigate in vivo response of cholinesterase(ChE) in anticholinesterase exposures,whereas the type constitution and property of the enzyme remained unclear.A type of ChE was purified from Daphnia magna using a three-step procedure,i.e.,Triton X-100 extraction,ammonium sulfate precipitation,and diethylaminoethyl(DEAE)-Sepharose Fast-Flow chromatography.According to sodium dodecyl sulfate polyacrylamide gel electrophoresis(SDS-PAGE),molecular mass of the purified ChE was estimated to be 84 kDa.Based on substrate studies,the purified enzyme preferred butyrylthiocholine iodide(BTCh) [with maximum velocity(Vmax)/Michaelis constant(Km)=8.428 L/(min·mg protein)] to acetylthiocholine iodide(ATCh) [with V max /Km =5.346 L/(min·mg protein)] as its substrate.Activity of the purified enzyme was suppressed by high concentrations of either ATCh or BTCh.Inhibitor studies showed that the purified enzyme was more sensitive towards inhibition by tetraisopropylpyrophosphoramide(iso-OMPA) than by 1,5-bis(4-allyldimethylammoniumphenyl) pentan-3-one dibromide(BW284C51).Result of the study suggested that the purified ChE was more like a type of pseudocholinesterase,and it also suggested that Daphnia magna contained multiple types of ChE in their bodies.