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. 2016 Jul 5;291(34):17754–17771. doi: 10.1074/jbc.M116.727107

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© 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

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FIGURE 3. — Purification and enzymatic characterization of recombinant LdTyrRS. A, purification of recombinant LdTyrRS protein on nickel-nitrilotriacetic acid affinity resin. M, molecular weight marker; Lane 1, uninduced cell lysate; lane 2, induced cell lysate; lane 3, rLdTyrRS. B, Western blotting analysis of the recombinant LdTyrRS protein using anti-His tag mouse antibody (1:3000). rTyrRS, purified rLdTyrRS. C, immunoblotting analysis of the Leishmania promastigote (P) and amastigote (A) cell lysate (∼40 μg) with the anti-LdTyrRS antibody. D, time course of tRNA^Tyr aminoacylation by recombinant LdTyrRS. Reactions were performed with l-tyrosine and tRNA^Tyr as the substrates. The data shows an average of three experiments performed in duplicate ± S.D. E and F, aminoacylation kinetics of LdTyrRS as a function of l-tyrosine concentration (E) or tRNA^Tyr concentration (F). The results represent mean ± S.D. with n = 3.