The kinetics of haemoglobin. II. The velocity with which oxygen dissociates from its combination with haemoglobin.
The kinetics of haemoglobin. II. The velocity with which oxygen dissociates from its combination with haemoglobin.
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DOI：
10.1098/rspa.1923.0117
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发表时间：
1923-10-01
期刊：
影响因子：
--
通讯作者：
Roughton, FJW
中科院分区：
文献类型：
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作者：
Hartridge, H;Roughton, FJW
对于生理学家和物理化学家来说，氧与球蛋白结合和解离的速度是一个相当感兴趣的问题；前者是因为这种化合物在呼吸中执行的非常重要的部分，而后者是因为球蛋白是一个几乎唯一的大的复杂蛋白质分子，它与气体结合，显然不是通过吸附，而是以一种简单的化学方式，由质量作用定律定义。在我们的主要实验研究可以开始之前，有几个初步问题是我们必须解决的。我们的第一个主要问题是找到一种非常突然的方法来破坏存在于溶液中的氧、氢球蛋白和氧气球蛋白之间的化学平衡。打破平衡所需的时间必须比系统恢复化学平衡所需的时间短得多。在反应CO+O2Hb⇄O，+COHb的情况下，通过将溶液暴露在强大的光束中解决了这个问题；后者导致了一个新的平衡位置被占据，这可能会被光束的突然中断瞬间打乱。于是，这个系统又回到了暗平衡的位置。为了使溶液的所有部分都经过所产生的反应的相同阶段，与恢复平衡所需的时间相比，打破平衡所需的时间必须可以忽略不计。不幸的是，在目前的情况下，没有向我们开放类似的方法，因为反应O2+Hb⇄O2Hb即使有影响，也不明显地受到强大的光束的影响。近几年来，Barcroft和他的同事们已经非常彻底地研究了这个体系的平衡所依赖的因素，主要的因素是温度、氢离子浓度和溶液的含盐量。计算表明，即使我们有一些非常突然的方法来改变这些因素中的一个或多个，与可用的定量方法的实验误差相比，系统的移位量也太小了。因此，我们采取的计划是准备一种球蛋白溶液I和另一种溶液II，以便如果I和II非常迅速但完全混合，则混合后的溶液立即不处于化学平衡状态，而是在一段时间后达到平衡，与混合过程所需的时间相比，该间隔时间较长。因此，作为一个例子，我们可以提到，在研究凝血球蛋白的氧化速率时，溶液I由稀释的还原的凝血球蛋白组成，而溶液II由含有足够溶解的O2的水组成，以与I的凝血球蛋白结合。在I和II非常快速地混合之后的瞬间，凝血球蛋白仍被部分还原，并通过稍后描述的方法测量其随后变得完全氧化的速率。为了这一计划的成功，有必要设计一种特殊类型的混合设备，其描述和测试已经在我们以前的一篇论文中进行了描述(2)。
The velocity with which oxygen combines with, and is dissociated from, hæmoglobin is a matter of considerable interest both to the physiologist and the physical chemist; to the former because of the all-important part performed by this compound in respiration, and to the latter because hæmoglobin is an almost unique example of a large complex protein molecule which combines with gases, apparently not by adsorption, but in a simple chemical manner defined by the laws of mass action. There were several preliminary problems which it was necessary for us to solve, before our main experimental investigation could be commenced. Our first major problem was to find some very sudden method of upsetting the chemical equilibrium subsisting between oxygen, hæmoglobin and oxyhæmoglobin in solution. The time taken to upset the equilibrium must be very much shorter than the time taken by the system to regain chemical equilibrium. This problem in the case of the reaction CO+O2Hb⇄O,+COHb was solved by exposing the solution to a powerful beam of light; the latter caused a new position of equilibrium to be taken up, and this could be instantaneously upset by a sudden interruption of the beam of light. The system thereupon returned to its position of dark equilibrium. In order that all parts of the solution shall be passing through the same stages of the resulting reaction it is necessary that the time taken for the equilibrium to be disturbed be of negligible duration compared with that taken for equilibrium to be regained. Unfortunately, a similar method was not open to us in the present case, for the reaction O2+ Hb ⇄ O2Hb is not appreciably, if at all, affected by a powerful beam of light. The factors upon which the equilibrium of this system depends have been very thoroughly studied by Barcroft and his co-workers in recent years (18) ; the principal ones are the temperature, the hydrogen ion concentration and salt content of the solution. Calculations showed that even if we had some very sudden method of changing one or more of these factors, the amount by which the system would be displaced would be too small compared with the experimental error of the quantitative methods available. The plan which we have therefore adopted was to prepare a hæmoglobin solution I and another solution II such that if I and II are very rapidly,but completely, mixed, the solution immediately after mixing is not in chemical equilibrium, but reaches equilibrium after an interval of time which is long in comparison with the time taken up by the process of mixing. Thus as an example we may mention that, in studying the rate of oxidation of hæmoglobin, solution I consisted of dilute reduced hæmoglobin, whilst solution II consisted of water containing sufficient dissolved O2to combine with the hæmoglobin of I. At the instant after the very rapid mixing of I and II the hæmoglobin is still partially reduced, and by methods to be described later the rate at which it subsequently becomes fully oxidised is measured. For this plan to be successful it was necessary to devise a special type of mixing apparatus, the description and testing of which have already been described in one of our previous papers (2).