Brett Ian C, Johansson Bert E
Department of Pediatrics, Division of Critical Care Medicine, Weill Medical College of Cornell University, 525 East 68th St., New York, NY 10021, USA.
J Biochem. 2006 Mar;139(3):439-47. doi: 10.1093/jb/mvj051.
Enzymatic kinetic parameters of influenza A virus N1 neuraminidases (NA) chromatographically purified from several vaccine candidate strains were tested. With ionic strength held constant, Ca2+ or Mg2+ increased the initial rate of enzymatic activity. The 1934 and 1943 strains had statistically significant highest initial velocities, V(max)/K(m) and V(max). There were no significant differences among the influenza virus strains from 1947 to 1991. Measured K(m) for the 1943 strain (6.2 x 10(-5) M) was significantly higher than other strains (3.1-4.7 x 10(-5) M). V(max)/K(m) varied from 0.78 M(-1) s(-1) to 0.91 M(-1) s(-1) and V(max) varied from 3.0 s(-1) to 5.5 s(-1) before the addition of a divalent cation and increased approximately 2-fold for each of these kinetic parameters for each strain after the addition of exogenous Ca2+ or Mg2+. Dialysis reduced the initial velocity and immunogenicity of each strain with significant differences found among strains. Enzymatic activity and immunogenicity were partially restored by the addition of exogenous Ca2+. Nucleic acid sequence analysis could not predict these differences. Selection of vaccine strains must include analysis of antigenic changes, but also enzymatic studies and determination of the requirement of divalent cations to maintain immunogenicity and activity during production.
对从几种候选疫苗株中通过色谱法纯化得到的甲型流感病毒N1神经氨酸酶（NA）的酶动力学参数进行了测试。在离子强度保持恒定的情况下，Ca2+或Mg2+提高了酶活性的初始速率。1934株和1943株具有统计学上显著最高的初始速度、V(max)/K(m)和V(max)。1947年至1991年的流感病毒株之间没有显著差异。1943株测得的K(m)（6.2×10(-5)M）显著高于其他株（3.1 - 4.7×10(-5)M）。在添加二价阳离子之前，V(max)/K(m)在0.78M(-1)s(-1)至0.91M(-1)s(-1)之间变化，V(max)在3.0s(-1)至5.5s(-1)之间变化，添加外源性Ca2+或Mg2+后，每种菌株的这些动力学参数均增加了约2倍。透析降低了每种菌株的初始速度和免疫原性，各菌株之间存在显著差异。添加外源性Ca2+可部分恢复酶活性和免疫原性。核酸序列分析无法预测这些差异。疫苗株的选择必须包括对抗抗原变化的分析，还包括酶学研究以及确定在生产过程中维持免疫原性和活性所需的二价阳离子。
