📚 Factors that Affect Enzyme Action | 影响酶作用的因素
Enzymes are globular proteins that act as biological catalysts, lowering activation energy and accelerating reactions without being consumed. Their activity is highly sensitive to environmental conditions and to the concentrations of molecules that bind to them. For Cambridge A-Level Biology, you must explain how temperature, pH, enzyme concentration, substrate concentration, inhibitors, cofactors and immobilisation influence the rate of an enzyme-catalysed reaction, and you must link these effects to protein structure and the active site.
酶是球状蛋白质，作为生物催化剂，可降低活化能并加速反应，而自身不被消耗。酶活性对周围环境以及与酶结合的分子浓度高度敏感。在剑桥 A-Level 生物学中，你必须解释温度、pH、酶浓度、底物浓度、抑制剂、辅因子和固定化如何影响酶促反应速率，并将这些影响与蛋白质结构和活性部位联系起来。
1. Enzyme Structure and the Active Site | 酶的结构与活性部位
An enzyme is a globular protein with a precise three-dimensional shape held together by hydrogen bonds, ionic bonds, hydrophobic interactions and, in some cases, disulfide bridges. Within this structure, a small region called the active site is formed by a few amino acid side chains. The active site is usually a groove or cleft on the enzyme surface, and its shape, charge and hydrophobic character are complementary to the substrate.
酶是球状蛋白质，其精确的三维形状由氢键、离子键、疏水相互作用以及某些情况下的二硫键维持。在这一结构中，由少数氨基酸侧链形成一个小区域，称为活性部位。活性部位通常是酶表面的凹槽或裂隙，其形状、电荷和疏水特性与底物互补。
The active site is not the whole enzyme; it is only a small part of the molecule. The specific folding of the polypeptide chain brings together amino acids that may be far apart in the primary sequence. This means that any factor that changes the shape, charge or flexibility of the active site can reduce the formation of enzyme-substrate complexes and therefore reduce the rate of reaction. The rate of an enzyme-catalysed reaction is usually measured as the initial rate, before substrate is depleted or product accumulates.
活性部位不是整个酶，而只是分子的一小部分。多肽链的特定折叠使一级结构中可能相距很远的氨基酸聚集在一起。这意味着任何改变活性部位形状、电荷或柔韧性的因素都会减少酶-底物复合物的形成，从而降低反应速率。酶促反应速率通常以初速率测定，即在底物被消耗或产物积累之前测定。
Enzymes lower activation energy by providing an alternative reaction pathway. They do not change the equilibrium position of a reversible reaction, and they are not used up in the reaction. After catalysis, the enzyme is released unchanged and can bind another substrate molecule.
酶通过提供替代反应途径来降低活化能。它们不改变可逆反应的平衡位置，也不在反应中被消耗。催化后，酶不变地被释放，并可结合另一个底物分子。
2. Models of Enzyme Action: Lock-and-Key and Induced Fit | 酶作用模型：锁钥与诱导契合
The lock-and-key model proposes that the active site has a rigid shape that is exactly complementary to the substrate, like a key fitting a lock. This model explains why enzymes are highly specific: only a substrate with the correct shape can fit into the active site and form an enzyme-substrate complex.
锁钥模型提出，活性部位具有与底物完全互补的刚性形状，就像钥匙插入锁中。该模型解释了酶为何具有高度特异性：只有形状正确的底物才能进入活性部位并形成酶-底物复合物。
The induced-fit model is now more widely accepted. It suggests that the active site is not completely rigid. When the substrate binds, the active site changes shape slightly so that it fits more closely around the substrate. This change can strain chemical bonds in the substrate, making them easier to break and lowering the activation energy further.
诱导契合模型目前被更广泛接受。它认为活性部位并非完全刚性。当底物结合时，活性部位会略微改变形状，使其更紧密地包裹底物。这种变化可以拉扯底物中的化学键，使其更容易断裂，从而进一步降低活化能。
Induced fit also helps explain why some molecules can act as competitive inhibitors. If a molecule is similar in shape to the substrate, it may bind to the active site and prevent the substrate from binding. Similarly, molecules that bind away from the active site can change the shape of the active site, which is the basis of non-competitive inhibition.
诱导契合也有助于解释为何某些分子可作为竞争性抑制剂。如果某分子在形状上与底物相似，它可能结合到活性部位并阻止底物结合。同样，结合在活性部位以外区域的分子可以改变活性部位形状，这就是非竞争性抑制的基础。
3. Measuring the Rate of Enzyme-Catalysed Reactions | 酶促反应速率的测定
The rate of an enzyme-catalysed reaction is the amount of substrate used or product formed per unit time. In experiments, it is common to measure the initial rate, because as the reaction proceeds the substrate concentration falls and product concentration rises. These changes can slow the reaction, so the initial rate gives the clearest indication of the enzyme's activity under the chosen conditions.
酶促反应速率是单位时间内底物消耗量或产物生成量。实验中通常测定初速率，因为随着反应进行，底物浓度下降、产物浓度上升。这些变化会减慢反应，因此初速率能最清楚地反映所选条件下酶的活性。
Common methods of measurement include measuring the volume of gas produced using a gas syringe or inverted burette, measuring the decrease in mass using a balance, measuring colour change using a colorimeter, and measuring pH change using a pH meter. The rate is found from the gradient of a tangent drawn to the curve at the start of the reaction.
常用测定方法包括：用气体注射器或倒置滴定管测量气体体积，用天平测量质量减少，用色度计测量颜色变化，用 pH 计测量 pH 变化。速率可从反应开始时曲线切线的斜率求出。
For accuracy, all variables other than the one being investigated must be controlled. This includes temperature, pH, substrate concentration, enzyme concentration and the volume of the reaction mixture. When plotting a graph, the independent variable is placed on the x-axis and the dependent variable on the y-axis. A steep initial gradient indicates a fast reaction; a plateau indicates that the rate has reached a maximum.
为保证准确性，除被研究的变量外，其他变量都必须控制，包括温度、pH、底物浓度、酶浓度和反应混合物体积。作图时，自变量放在 x 轴，因变量放在 y 轴。初始斜率陡表示反应快；平台表示速率达到最大值。
4. Effect of Temperature | 温度的影响
As temperature increases from a low value, the kinetic energy of enzyme and substrate molecules increases. This leads to more frequent and more energetic collisions, so more enzyme-substrate complexes form and the rate of reaction increases. Over a limited range, the rate approximately doubles for every 10 °C rise; this is described by the temperature coefficient, Q₁₀.
当温度从较低值升高时，酶和底物分子的动能增加，导致碰撞更频繁、更有能量，因此形成更多酶-底物复合物，反应速率增加。在有限范围内，温度每升高 10 °C，速率大约翻倍；这用温度系数 Q₁₀ 描述。
However, above the optimum temperature, the rate falls sharply. The increased kinetic energy causes hydrogen bonds and ionic bonds that maintain the tertiary structure to break. The active site loses its specific shape, so the substrate can no longer bind effectively. The enzyme is said to be denatured. Denaturation is usually irreversible because the precise three-dimensional folding of the polypeptide chain cannot easily be restored.
然而，超过最适温度后，速率急剧下降。增加的动能导致维持三级结构的氢键和离子键断裂。活性部位失去特定形状，底物不再能有效结合。这种酶被称为变性。变性通常是不可逆的，因为多肽链精确的三维折叠难以恢复。
The optimum temperature varies between enzymes and organisms. Human enzymes usually have an optimum near 37 °C. Enzymes from thermophilic bacteria may have optima above 70 °C, while enzymes from cold-adapted organisms may work best at low temperatures. This is important in biotechnology and in understanding how organisms survive in extreme environments.
最适温度因酶和生物体而异。人体酶的最适温度通常接近 37 °C。嗜热细菌的酶最适温度可能高于 70 °C，而适应寒冷的生物体的酶可能在低温下活性最佳。这在生物技术和理解生物如何生存在极端环境中很重要。
| Temperature range | Molecular explanation | Rate change |
|---|---|---|
| Below optimum | More kinetic energy, more effective collisions, more ES complexes | Increases |
| At optimum | Maximum frequency of successful collisions without denaturation | Maximum |
| Above optimum | Hydrogen and ionic bonds break, tertiary structure changes, active site denatures | Decreases sharply |
5. Effect of pH | pH 的影响
pH is a measure of the hydrogen ion concentration, [H⁺]. A low pH means a high concentration of H⁺ ions, while a high pH means a low concentration of H⁺ ions and a higher concentration of OH⁻ ions. Changes in pH alter the charges on amino acid side chains in the active site, especially -COOH and -NH₂ groups. This can disrupt hydrogen bonds and ionic bonds that hold the tertiary structure together.
pH 是氢离子浓度 [H⁺] 的度量。低 pH 意味着 H⁺ 离子浓度高，高 pH 意味着 H⁺ 离子浓度低而 OH⁻ 离子浓度较高。pH 变化会改变活性部位中氨基酸侧链的电荷，尤其是 -COOH 和 -NH₂ 基团。这会破坏维持三级结构的氢键和离子键。
If the charge on the active site changes, the substrate may no longer bind correctly, so fewer enzyme-substrate complexes form and the rate decreases. At extreme pH values, the enzyme becomes denatured because the tertiary structure is permanently disrupted. This is why enzymes have a narrow optimum pH range and why pH must be controlled in experiments using buffers.
如果活性部位电荷改变，底物可能不再正确结合，因此形成的酶-底物复合物减少，速率下降。在极端 pH 下，酶会变性，因为三级结构被永久破坏。这就是酶具有狭窄最适 pH 范围的原因，也是实验中必须使用缓冲液控制 pH 的原因。
Different enzymes have different optimum pH values because their active sites contain different amino acid side chains. Pepsin, which works in the stomach, has an optimum pH of about 2. Trypsin, which works in the small intestine, has an optimum pH of about 8. Salivary amylase has an optimum pH of about 6.8. These values reflect the environment in which each enzyme normally functions.
不同酶具有不同的最适 pH，因为其活性部位含有不同的氨基酸侧链。胃中工作的胃蛋白酶最适 pH 约为 2。小肠中工作的胰蛋白酶最适 pH 约为 8。唾液淀粉酶最适 pH 约为 6.8。这些数值反映了每种酶正常发挥作用的环境。
| Enzyme | Optimum pH | Location |
|---|---|---|
| Pepsin | About 2 | Stomach |
| Salivary amylase | About 6.8 | Mouth |
| Trypsin | About 8 | Small intestine |
6. Effect of Enzyme Concentration | 酶浓度的影响
When substrate concentration is not limiting, the rate of reaction is directly proportional to enzyme concentration. This is because doubling the number of enzyme molecules doubles the number of active sites available. With more active sites, more enzyme-substrate complexes can form per unit time, so the rate increases.
当底物浓度不是限制因素时，反应速率与酶浓度成正比。这是因为酶分子数量加倍，可利用的活性部位数量也加倍。活性部位越多，单位时间内可形成更多酶-底物复合物，因此速率增加。
If substrate concentration is limited, increasing enzyme concentration will only increase the rate until all substrate molecules are being used as fast as possible. After that point, the graph levels off because there is not enough substrate to occupy all the active sites. This is an important experimental point: to study the effect of enzyme concentration, substrate must be in excess.
如果底物浓度有限，增加酶浓度只能使速率增加到所有底物分子尽可能快地被利用为止。此后曲线趋于平缓，因为没有足够底物占据所有活性部位。这是一个重要的实验要点：研究酶浓度影响时，底物必须过量。
The graph of initial rate against enzyme concentration is usually a straight line through the origin, and the gradient represents the catalytic efficiency under those conditions. If the line does not pass through the origin, it may indicate that the enzyme preparation already contained some product, or that there is a background reaction.
初速率对酶浓度作图通常是通过原点的直线，其斜率代表该条件下的催化效率。如果直线不通过原点，可能表明酶制剂中已含有一些产物，或存在背景反应。
7. Effect of Substrate Concentration and Vmax/Km | 底物浓度的影响与 Vmax/Km
At low substrate concentrations, the rate of reaction is directly proportional to substrate concentration. Most active sites are empty, so increasing substrate concentration increases the chance of collision and the number of enzyme-substrate complexes formed. This is described as first-order kinetics with respect to substrate.
在低底物浓度下，反应速率与底物浓度成正比。大多数活性部位是空的，因此增加底物浓度会增加碰撞机会和形成的酶-底物复合物数量。这被称为对底物的一级动力学。
As substrate concentration increases further, the rate increases more slowly. Eventually, all active sites become occupied at any instant. The enzyme is said to be saturated. At this point, adding more substrate does not increase the rate because there are no free active sites available. The maximum rate is called Vmax, and the reaction is zero-order with respect to substrate.
随着底物浓度进一步增加，速率增加变慢。最终，所有活性部位在任一时刻都被占据。酶被称为饱和。此时，再加入更多底物也不会增加速率，因为没有可用的游离活性部位。最大速率称为 Vmax，反应对底物
更多咨询请联系16621398022（同微信）
或截图后用「扫一扫 → 相册」选择本图