(Bio)chemistry: TestAS practice testlets

Enzymes are biological catalysts, mostly proteins, that speed up chemical reactions by lowering their activation energy without being used up. The molecule an enzyme acts on, the substrate, binds to a specific region, the active site, whose shape and chemical properties fit the substrate. After the reaction, the product is released and the enzyme can bind the next substrate molecule.

The rate of an enzyme reaction depends on the substrate concentration. At low concentrations, the rate rises almost in proportion to the concentration, because most active sites are free. At high concentrations, nearly all active sites are occupied, and the rate approaches a maximum, Vmax, which cannot be exceeded however much substrate is added; the enzyme is saturated. This behaviour is described by the Michaelis-Menten equation. The Michaelis constant Km is the substrate concentration at which the rate is half of Vmax. A low Km means that the enzyme reaches half its maximum rate at a low substrate concentration, which indicates a high affinity for the substrate.

v = Vmax × [S] ÷ (Km + [S])

Inhibitors reduce enzyme activity. A competitive inhibitor resembles the substrate and competes for the active site. Its effect can be overcome by adding more substrate, so Vmax is unchanged, but more substrate is needed to reach half of Vmax: the apparent Km increases. A non-competitive inhibitor binds elsewhere on the enzyme and changes its shape, so that some enzyme molecules cannot work at all. Adding more substrate does not help; Vmax falls, while Km stays the same. Many drugs are enzyme inhibitors, and cells regulate metabolic pathways by inhibiting key enzymes.

Enzymes also depend on their environment. Activity rises with temperature up to an optimum, which is about 37 °C for most human enzymes, and falls steeply above it as the protein denatures. Each enzyme also has a pH optimum: pepsin works best at about pH 2 in the stomach, trypsin at about pH 8 in the small intestine.

Knowing Km and Vmax allows the rate at any substrate concentration to be calculated with the equation. Biochemists also use these values to compare enzymes from different organisms, to identify how a drug inhibits its target and to design industrial processes in which enzymes are used, for example in detergents or food production.

An enzyme was studied in the laboratory at 37 °C and pH 7. Its Vmax is 100 µmol of product per minute, and its Km is 2 mmol/L.

Chemists count particles in moles. One mole contains about 6.022 × 10²³ particles, a number chosen so that the mass of one mole of a substance in grams equals its relative molecular mass. The molar mass of glucose (C6H12O6) is therefore about 180 g/mol, and that of sodium chloride (NaCl) about 58.4 g/mol. The amount of a substance in moles is its mass divided by its molar mass.

The concentration of a solution is most often given as molarity, the number of moles of solute per litre of solution, with the unit mol/L, often written M. A 1 M glucose solution contains 180 g of glucose per litre. To prepare a solution, the required amount of substance is calculated as concentration times volume, converted into grams, dissolved in part of the solvent and then made up to the final volume.

n = m ÷ M c = n ÷ V m = c × V × M

Concentrations are also given as mass per volume (for example g/L or mg/mL) or as percentages. In biology and medicine, "0.9% sodium chloride", physiological saline, means 0.9 g of NaCl per 100 mL, or 9 g per litre. Its molarity is 9 ÷ 58.4, about 0.154 mol/L. Because each NaCl unit releases two ions in water, the solution contains about 0.308 mol/L of dissolved particles, which is close to the osmotic concentration of blood plasma.

Dilution is one of the most common laboratory tasks. When a solution is diluted, the amount of solute stays the same while the volume increases, so the product of concentration and volume is constant. For large dilution factors, laboratories use serial dilutions: a series of steps, each diluting the previous solution by the same factor. Three successive 1 : 10 dilutions give a total dilution of 1 : 1,000. Serial dilutions are used, for example, to count bacteria, since a sample with millions of cells per millilitre must be diluted before colonies can be counted on a plate.

c1 × V1 = c2 × V2

Careful work also requires attention to units. Volumes in millilitres must be converted into litres before they are multiplied by a concentration in mol/L, and masses in milligrams into grams. A quick plausibility check, such as asking whether a few grams of substance for a few hundred millilitres of solution is reasonable, catches many errors before a solution is prepared.