International unit of enzyme activity

The reader should note that U had been reserved by chemists for the international unit of enzyme activity. However, U is often used to mean a unit of TSH concentration as defined by the WHO. To avoid confusion with enzyme activity units, lU and int units are used in this chapter to refer to the units of TSH concentration. 

In many situations, the actual molar amount of the enzyme is not known. However, its amount can be expressed in terms of the activity observed. The International Commission on Enzymes defines One International Unit of enzyme as the amount that catalyzes the formation of one micromole of product in one minute. (Because enzymes are very sensitive to factors such as pH, temperature, and ionic strength, the conditions of assay must be specified.) Another definition for units of enzyme activity is the katal. One katal is that amount of enzyme catalyzing the conversion of one mole of substrate to product in one second. Thus, one katal equals 6X10 international units. 

This corresponds to the recommendations given in 1959 by a joint committee of the Clinical Chemistry Commission of IUPAC (International Union of Pure and Applied Chemistry) and the Enzyme Commission of IUB (International Union of Biochemistry). Thus, one unit of enzyme activity should be defined as that amount of enzyme which catalyzes the conversion of one micromole of substrate per minute under defined conditions (W9). 

Because of the difficulties in measuring the amount of enzyme in the conventional units of mass or molar concentration, the accepted unit of enzyme activity is defined in terms of reaction rate. The International Unit (IU) is defined as that amount of enzyme which will result in the conversion of 1 /nmol of substrate to product in 1 minute under specified conditions. The SI unit of activity, which is becoming more acceptable, is the katal and is defined as that amount of enzyme which will result in the conversion of 1 mol of substrate to product in 1 second. A convenient sub-unit is the nanokatal, which is equal to 0.06 International Units. 

International Unit (IU) Ratal (kat), the true SI unit of enzyme activity Specific activity that amount of enzyme protein which brings about the conversion of 1 (Imol of substrate to product per minute under stated conditions that amount of enzyme protein which brings about the conversion of 1 mole of substrate to product per second under stated conditions. enzyme activity (as IU or kat) per mg of total protein. This is a useful measure of the purity of an enzyme preparation... 

By international agreement, 1.0 unit of enzyme activity is defined as the amount of enzyme causing transformation of 1.0 gmol of substrate per minute at 25 °C under optimal conditions of measurement. The term activity refers to the total units of enzyme in a solution. The specific activity is the number of enzyme units per milligram of total protein (Fig. 3-23). The specific activity is a measure of enzyme purity it increases during purification of an enzyme and becomes maximal and constant when the enzyme is pure (Table 3-5). 

The actual molar concentration of an enzyme in a cell-free extract or purified preparation is seldom known. Only if the enzyme is available in a pure crystalline form, carefully weighed, and dissolved in a solvent can the actual molar concentration be accurately known. It is, however, possible to develop a precise and accurate assay for enzyme activity. Consequently, the amount of a specific enzyme present in solution is most often expressed in units of activity. Three units are in common use, the international unit (IU), the katal, and specific activity. The International Union of Biochemistry Commission on Enzymes has recommended the use of a standard unit, the international unit, or just unit, of enzyme activity. One IU of enzyme corresponds to the amount that catalyzes the transformation of 1 p,mole of substrate to product per minute under specified conditions of pH, temperature, ionic strength, and substrate concentration. If a solution containing... 

The spectrophotometer measures and displays the increase in absorbance at 410 nm as a function of time (AA/At). Whether the output from the instrument is in the form of a strip chart or is collected by a computer, the reaction velocities are usually expressed in terms of change in concentration per unit time, or converted to specified units of enzyme activity. The International Unit (U) for enzyme activity is defined as the amount of enzyme that transforms 1 pmol substrate to product in 1 min under specified assay conditions. The SI unit for activity is the katal, which is defined as the amount of enzyme that transforms 1 mol substrate per second under specified conditions. Thus 1 U = 16.7 nkatal. To convert slopes AA/At values) to velocities (v), the following equation is used ... 

The esterification activity of Lipozyme IM-20 was measured according to the method described by Langone and Sant Anna (9), which determines the consumption rate of fatty acid at 60°C in a reaction system containing glycerol, lauric acid, and a given amount of the commercial enzyme preparation. One international unit of esterification activity is the quantity of enzyme that consumes 1 pmol of lauric acid/min under the reaction conditions. The enzyme used has an esterification activity of 20 IU/g. 

The Jhtemationai Union of Biochemistry (TUB) was officially founded in 1953 by an initiative of the British Biochemical Society. At ibis time enzyme standardization was in a chaotic slate, owing to the multiplicity of arbitrarily defined units of enzyme activity and the ilt-dtfned nomenclature. In 1955, the 1UB International Commission on Enzymes was created. This led to an improved enzyme nomoidature, which has been used since 1961, and the definition of the International Unit (LU.) of enzyme activity. 

Calculation One unit of enzyme activity (FIP Unit) is defined as that quantity of a standard lipase preparation (Fungi Lipase-International FIP Standard) that liberates the equivalent of 1 (imol of fatty acid per minute from the Substrate Emulsion under the described assay conditions. The specific activity is expressed in international FIP units per milligram of the Sample Preparation. 

The routine unit of enzyme activity has been the international unit (I.U.), namely xmoles P formed (or S consumed) per minute. The specific activity of an enzyme preparation is the number of xmoles P formed (or S consumed) per minute per milligram of protein (clearly this will be very low in a crude cell extract and have a maximal value for a pure preparation of the enzyme). If the molecular mass is known, the specific activity of a pure enzyme measured in saturating (Fmax conditions) can be used to calculate the turnover number (or molecular activity ) of an enzyme, namely the number of P molecules formed (or S molecules transformed) per molecule of enzyme per second (units sec- ). If we recall that the maximal velocity (Fmax) equals k+2 (sec " ) [ET], we can see that the molecular activity equals k+2 (sec -1), that is, fal (sec-1). The katal is the S.I. unit of enzyme activity (moles substrate transformed sec -I) from whence come the corresponding units for specific activity (katal kilogram-1) and molar activity (katal per mole of enzyme). 

A new unit of enzyme activity, the katal, has been proposed. One katal is that amount of enzyme that catalyzes the conversion of one mole of substrate per second. Thus, one International Unit 1/60/ikatal 16.67 nkatal. One katal = 6 X lO Units. Specific activity can be expressed as katals/kg protein or p.katals/mg protein. The molar activity of an enzyme is the katals/mole protein with units of sec . 

To standardize how enzyme activities are expressed, the Enzyme Commission of the lUB proposed that the unit of enzyme activity be defined as the quantity of enzyme that catalyzes the reaction of 1 pmol of substrate per minute and that this unit be termed the international unit (U). Catalytic concentration is to be expressed in terms of U/L or kU/L, whichever gives the more convenient numerical value. In this chapter, the symbol U is used to denote the international unit. In those instances in which there is some uncertainty about the exact nature of the substrate or when there is difficulty in calculating the number of micromoles reacting (as with macromolecules such as starch, protein, and complex lipids), the unit is to be expressed in terms of the chemical group or residue measured in following the reaction (e.g., glucose units, or amino acid units formed). 

Nomenclature Committee of the International Union of Biochemistry (NC-IUB) (1979) Units of enzyme activity, recommendations 1979. Eur JBiochem 97 319-320... 

Enzymes themselves can be analyzed by measuring the amount of substrate transformed in a given time or the product that is produced in a given time. The substrate, houldJ2eJn ccess so that the reaction rate depends oiily on the enzyme concentration. The results are expressed as international units of enzyme. For example, the activity of a glucose oxidase preparation can be determined by measuring manometrically or amperometrically the number of micromoles of oxygen consumed per minute. On the other hand, the use of enzymes to develop specific procedures for the determination of substrates, particularly in clinical chemistry, has proved to be extremely useful. In this case, the enzyme concentration is in excess so the reaction rate is dependent on the substrate concentration. 

Enzyme activity is expressed in units of activity. The Enzyme Commission of the International Union of Biochemistry recommends to express it in international units (lU), defining 1 lU as the amount of an enzyme that catalyzes the transformation of 1 pmol of substrate per minute under standard conditions of temperature, optimal pH, and optimal substrate concentration (International Union of Biochemistry). Later on, in 1972, the Commission on Biochemical Nomenclature recommended that, in order to adhere to SI units, reaction rates should be expressed in moles per second and the katal was proposed as the new unit of enzyme activity, defining it as the catalytic activity that will raise the rate of reaction by 1 mol/second in a specified assay system (Anonymous 1979). This latter definition, although recommended, has some practical drawbacks. The magnitude of the katal is so big that usual enzyme activities expressed in katals are extremely small numbers that are hard to appreciate the definition, on the other hand, is rather vague with respect to the conditions in which the assay should be performed. In practice, even though in some journals the use of the katal is mandatory, there is reluctance to use it and the former lU is still more widely used. 

As opposite to parameter K (or Keq) and kcat, V is not a fundamental property of the enzyme since it depends on its concentration as indicated by Eq. 3.5. This has to be taken into consideration when determining the kinetic parameters. The catalytic rate constant (kcat) is a fundamental property of the enzyme that can be expressed in different ways and in different units, according to how e is expressed (moles gL UL ). If e is expressed in moles L , kcat has dimension of T (known as turnover number). This requires the knowledge of the molecular weight and the specific activity and number of active centers of the enzyme. Sometimes this information is not available so that kcat is expressed in dimensions of M T (mass of substrate converted per unit time and unit of enzyme activity). If U is expressed in international units (lU), then kcat reduces to a dimensionless value of 1, which is to say that it is equivalent to V. 

The intrinsic variable expressed usually as international units (U) of enzyme activity per milligram protein... 

A new international unit has been recommended. The katal (kat) is defined as the amount of enzyme activity that transforms 1 mole of substrate per second. Activities can also be given in millikatals (mkat), microkatals (fxkat), nanokatals (nkat), etc. Both specific activity and turnover number can also be expressed in these units. 

In most preparations the actual molar concentration of enzyme is unknown. Consequently, the amount of enzyme present can be expressed only in terms of its activity. In order to standardize the reporting of enzyme activities, the Commission on Enzymes of the International Union of Biochemistry has defined a standard unit ... 

Because of the nonstandard units associated with the I.U. system for defining enzyme concentrations, an equivalent International system of units (Systeme International) (S.I.) unit has been defined, and is called the katal. One katal (kat) of enzyme activity is that quantity that will consume 1-mol substrate/s 1 pkat = 60 I.U. 

The base unit katal (symbol kat), mol/sec, is the catalytic amount of any catalyst, including enzymes, that catalyzes a reaction rate of 1 mol per second in an assay system." The kind of quantity measured is identified as catalytic amount. There is a constant relationship between the international unit (I pmol/min) and the katal (1 mol/sec) to convert, a y ue. in international units to nmol/sec, the value is multiplied by 16.67. Note, however, that dependence on reaction conditions applies to. SI units in the same way as to international units therefore data reported in the same units but obtained under different conditions may not be comparable. Replacement of the international unit for reporting enzyme activity is likely to be slow even units that antedated the international unit are sometimes used in clinical laboratory practice. (See Chapters 8 and 21 for further details on the expression of enzyme activity.)... 

The enzyme activity is defined as the moles of substrate or product converted in a reaction per time unit. The enzyme activity depends on the reaction conditions, which should be specified. In the International system (SI) the unit is the katal (i katal=1 mol/s) but a more practical and commonly used value is 1 enzyme unit (U) =l imol/min. One enzyme unit corresponds to 16.67 nanokatals [7]. 

There are no internationally standard assay methods for industrial enzymes and the definition of enzyme activity unit is also different for each enzyme. The activity of industrial enzymes is shown by various methods depending on manufacturers. For instance, commercial lipase activities are measured by the hydrolysis of olive oil under the various conditions and these figures are not comparable with each other. When customers apply these biocatalysts for chemical synthesis in organic solvents, these figure are sometimes reliable, and sometimes not. Users should not judge commercial enzymes based only on price and the activity shown in the table the manufacturer provides. Enzymes should be evaluated based on their practical performance under the conditions used. Most users of biotransformation are not experts in measuring enzyme activity, so the establishment of an assay method and practice are essential if one is to optimize the performance of enzymes. 

The activity of an enzyme preparation will vary from one source to another because the enzymes are usually not purified to 100% enzyme. That is, the percent enzyme will vary from one preparation to another. The activity of a given preparation is expressed in international units (I.U.). An international unit has been defined by the International Union of Biochemistry as the amount that will catalyze the transformation of one micromole of substrate per minute under defined conditions. The defined conditions will include temperature and pH. For example, a certain commercial preparation of glucose oxidase may have an activity of 30 units per milligram. Thus, for the determination of a substrate, a certain number of units of enzyme is taken. The specific activity is the units of enzyme per milligram of protein. Molecular activity is defined as units per molecule of enzyme, that is, it is the number of molecules of substrate transformed per minute per molecule of enzyme. The concentration of an enzyme in solution should be expressed as international units per milliliter or liter. 

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