n activity, the Kcat for apo and monomethylated substrate are 0.0033 min-1 and 0.015min-1, respectively, which also reflects a slightly faster reaction rate for the second step methylation. In the theoretical study of Rubisco LSMT, the potential energy barriers for the first and second methyl transfers calculated by MP2/6-31+G//MM were 21.4 kcal/mol and 19.6 kcal/mol. Therefore, both the computational and experimental results indicated a more efficient reaction catalyzed by PRMT1 than Rubisco LSMT. Although arginine is a weaker nucleophile than lysine, the methylation rate of the former was probably faster than the latter. This conclusion suggested that certain facilitating factors must be involved in PRMT1 active site to accelerate the reaction. In addition, although the potential energy barrier of the second methyl transfer was lower than the first, the value of experimental Michaelis constant Km reflects a relatively lower binding affinity of methylated substrates in the catalytic center than that of the apo substrate. Natural Bond Order analysis encoded in Gaussian 03 was performed to obtain Wiberg bond order diagram for further understanding the methyl transfer mechanism and explore why the second methyl transfer could be faster than the first one. In substrate arginine, guanidino cation is stabilized via efficient 26507655 resonance. Thus, the N atoms can be considered as between the sp2 and sp3 hybridization state. During reaction process, the bond order of NH2-CZ gradually decreased to 1 in TS, as shown in Sodium laureth sulfate chemical information proton Transfer Mechanism The experiment of solvent isotope effects suggested that no prior substrate deprotonation is required for PRMT1 catalysis. The following theoretical analysis based on QM calculations was performed to investigate the proton transfer 5 Catalytic Mechanism of PRMT1 doi: 10.1371/journal.pone.0072424.g002 6 Catalytic Mechanism of PRMT1 doi: 10.1371/journal.pone.0072424.g003 process involved in PRMT1 catalyzed arginine methylation. Confirmations extracted from the potential energy profile showed that the guanidino group was deprotonated immediately after methyl transfer, and the proton may transfer to the acid oxygen on E144. This result is 14557281 in accordance with the study on PRMT3, which proposed the proton transfers to E326, the counterpart of E144 in PRMT3. Thus, the important role of conserved glutamine in PRMT 7 Catalytic Mechanism of PRMT1 doi: 10.1371/journal.pone.0072424.g004 catalysis is revealed by these two theoretical investigations. NBO analysis was performed to explore the precedence relationship between methyl transfer and deprotonation. In the Wiberg bond order diagram, the concave shape of the line demonstrated that the formation of bond between OE2 and 2HH2 occurred after the formation of the bond between CE and NH2, or the proton transfer next to methyl transfer. We analyzed the evolution of electrostatic potential in the QM region for the first methyl transfer process to further understand the deprotonation of NH2. Charges on R54 and E153 remained constant during the reaction, whereas charges on Sub_R, AdoMet, and E144 showed apparent variations, suggesting these three residues were involved in reaction process, as demonstrated in Conclusion The present study revealed the mechanism of methyl transfer reaction catalyzed by arginine methyltransferase PRMT1 via theoretical computation. A model of PRMT1-substrate-cofactor complex was constructed, and a 30ns MD simulation was performed to ensure the stabi