Dawson Neal J, Katzenback Barbara A, Storey Kenneth B
Department of Biology and Institute of Biochemistry Carleton University, Ottawa, ON, Canada.
Department of Biology and Institute of Biochemistry Carleton University, Ottawa, ON, Canada.
Biochim Biophys Acta. 2015 Jan;1850(1):97-106. doi: 10.1016/j.bbagen.2014.10.003. Epub 2014 Oct 12.
The North American wood frog, Rana sylvatica, is able to overcome subzero conditions through overwintering in a frozen state. Freezing imposes ischemic and oxidative stress on cells as a result of cessation of blood flow. Superoxide dismutases (SODs) catalyze the redox reaction involving the dismutation of superoxide (O(2)(-)) to molecular oxygen and hydrogen peroxide.
The present study investigated the regulation of CuZnSOD and MnSOD kinetics as well as the transcript, protein and phosphorylation levels of purified enzyme from the muscle of control and frozen R. sylvatica.
CuZnSOD from frozen muscle showed a significantly higher V(max) (1.52 fold) in comparison to CuZnSOD from the muscle of control frogs. MnSOD from frozen muscle showed a significantly lower Km for O(2)(-) (0.66 fold) in comparison to CuZnSOD from control frogs. MnSOD from frozen frogs showed higher phosphorylation of serine (2.36 fold) and tyrosine (1.27 fold) residues in comparison to MnSOD from control animals. Susceptibility to digestion via thermolysin after incubation with increasing amount of urea (C(m)) was tested, resulting in no significant changes for CuZnSOD, whereas a significant change in MnSOD stability was observed between control (2.53 M urea) and frozen (2.92 M urea) frogs. Expressions of CuZnSOD and MnSOD were quantified at both mRNA and protein levels in frog muscle, but were not significantly different.
The physiological consequence of freeze-induced SOD modification appears to adjust SOD function in freezing frogs.
Augmented SOD activity may increase the ability of R. sylvatica to overcome oxidative stress associated with ischemia.
北美林蛙（Rana sylvatica）能够通过在冰冻状态下越冬来克服零下温度条件。由于血流停止，冷冻会给细胞带来缺血和氧化应激。超氧化物歧化酶（SOD）催化超氧化物（O₂⁻）歧化为分子氧和过氧化氢的氧化还原反应。
本研究调查了对照和冷冻的北美林蛙肌肉中铜锌超氧化物歧化酶（CuZnSOD）和锰超氧化物歧化酶（MnSOD）动力学的调节以及纯化酶的转录、蛋白质和磷酸化水平。
与对照蛙肌肉中的CuZnSOD相比，冷冻肌肉中的CuZnSOD显示出显著更高的Vmax（1.52倍）。与对照蛙的CuZnSOD相比，冷冻肌肉中的MnSOD对O₂⁻的Km显著更低（0.66倍）。与对照动物的MnSOD相比，冷冻蛙的MnSOD丝氨酸（2.36倍）和酪氨酸（1.27倍）残基磷酸化程度更高。在与增加量的尿素（C(m)）孵育后，测试了通过嗜热菌蛋白酶消化的敏感性，结果CuZnSOD没有显著变化，而在对照（2.53 M尿素）和冷冻（2.92 M尿素）蛙之间观察到MnSOD稳定性有显著变化。在蛙肌肉中对CuZnSOD和MnSOD的表达在mRNA和蛋白质水平上进行了定量，但没有显著差异。
冷冻诱导的SOD修饰的生理后果似乎是在冷冻蛙中调节SOD功能。
增强的SOD活性可能会增加北美林蛙克服与缺血相关的氧化应激的能力。
