Supplemental Lecture (97/02/06 update) by Stephen T. Abedon (firstname.lastname@example.org)
Typical name: substrate-ase:
- Chapter title: Enzymes
- A list of vocabulary words is found toward the end of this document
- The chemistry of life is generally completely dependent upon the control and catalysis of well defined reactions. Both control and catalysis is accomplished by structurally complex proteins called enzymes. Although convenient to consider enzymes as mere black boxes which serve to convert reactants to products, enzymes are actually dynamic molecules which interact with their environment in numerous ways in addition to simply catalyzing the conversion of reactant A into product B. In fact, it is environmental interaction plus response to environmental input which is key to the metabolic control effected by enzymes. Thus, within organisms, all sorts of anabolic and catabolic reactions can and do occur simultaneously.
- What products are utilized or are emphasized at any given moment is controlled by chemical inhibitors and enhancers of enzyme function. Also, many organic compounds lie at junction points between different biosynthetic pathways. Metabolism consequently is a multi-branched, reasonably well controlled "symphony" of enzymatically controlled and catalyzed chemical reactions.
- A protein catalyst employed by a cell to speed up metabolic reactions.
- Highly specific:
- Enzymes are highly specific in terms of what they act upon and what they do.
- This specificity and what the enzyme does is made possible by their structures.
Enzymes act on substrates.
They are usually named using the suffix -ase (e.g., synthetase or dehydrogenase). Without enzymes, life as we know it simply would not exist.
- An enzyme found and used within a cell.
- An enzyme secreted into the extracellular environment for use there, for example, in hydrolyzing polysaccharides to the monosaccharides which may then be taken up by cells and used.
- The chemical reactant acted upon by an enzyme.
- More generally, a substrate is a substance acted upon enzymatically by a microorganism and from which the microorganism derives nutrition.
- The word substrate is also used synonymously with substratum meaning, in addition to the above, the physical environment (possibly inert to enzyme action) upon which a microorganism lives, e.g., solid medium.
- Thus, glucose could be the substrate from which a microorganism derives carbon and energy. glucose is also acted upon by enzymes within the microorganism and thus is an enzymatic substrate . The microorganism may be living within an otherwise inert (i.e., its enzymes have no effect on it) mucous substrate attached to the lining of the large intestine.
- Sound confusing? Well, it is!
Following catalysis, the active site may be less hospitable to the presence of the altered molecule and thus product will be expelled from the active site.
- Physical location of enzyme activity:
- The physical position on an enzyme where a substrate(s) attaches.
- Often thought of as a pocket upon (or within) a globe-like (globular) protein molecule (see text figure 1045.1). Thus, to be acted upon, a substrate would first enter this active site pocket.
- The substrate's presence within this pocket might induce a conformational change in the protein (a.k.a., induced fit).
- Regardless, the active site would be shaped such that the substrate but not a differently shaped molecules would fit. Thus, the specificity of an enzyme is very often reflected in the shape of its active site.
- Stabilizes intermediate/product structures:
- Finally, amino acid R groups will project into the active site in such a manner that bonds in the substrate are rendered unstable and the bonds of the to-be-induced product molecule are more stable.
- Thus, catalysis to the favored product is induced to occur (see text figure 1045.2).
Note that an enzyme that requires a coenzyme for activity is known as an apoenzyme.
- A non-protein enzyme component, such as a metal ion or an organic compound, is called a cofactor (or coenzyme).
- Cofactors are found in active sites and play important roles in catalysis.
- Essential nutrients:
- Essential trace minerals are usually employed as cofactors.
- "Most trace elements required by living cells are probably used in some such way to activate cellular enzymes." (p. 106, Tortora et al., 1995)
- Coenzymes are cofactors that are organic compounds.
- Vitamins often serve as coenzymes.
Optimal enzyme activity
- A somewhat ambiguous term referring to the rate at which a reaction proceeds in the presence of a given amount of enzyme.
- This term is ambiguous because in practice one cannot be sure if an enzyme preparation is homogeneous. That is, given a pure preparation of a type of enzyme, can we be sure that all of the enzymes are functioning properly, i.e., active, or might some fraction be, for example, denatured? For simplicity, assume here that, in discussions of enzyme activity (below), we have a homogeneous enzyme population.
- Enzymes are both products of evolution and specifically adapted to certain uses in certain environments (often intracellular). A "cost" of such specificity is that activity tends to decline when environmental parameters differ from those found in the microenvironment within which the enzyme function evolved.
- Under conditions where such things as pH, temperature, or salt concentrations differ from those in which the enzyme function evolved, enzyme activity, minimally, can be expected to reversibly decline (see text figure 1045.3; these are the same things that are known to disrupt protein structure---denaturation).
- Buffering enhances enzyme activity:
- Cells are very good at buffering the intracellular environment such that it only vaguely resembles the extracellular environment.
- Note that of these three (temperature, osmolarity, and pH), a cell has the (very) least intracellular control over temperature, though multicelled organisms, such as mammals and birds, have considerably more control.
- Enzyme activity/function of substrate concentration:
- Enzymes are dependent on the presence of substrate to have activity. The more substrate present, the more substrate (absolutely) there is to be acted upon by a given amount of enzyme.
- That is, the rate at which substrate is acted upon is dependent entirely on the rate at which substrates molecules collide with enzymes, which in turn, all else being equal, is dependent almost entirely on substrate concentration.
- Diminishing returns:
- Eventually, though, there are diminishing returns meaning that the rate at which a substrate is processed can no longer be approximated as a linear function of substrate concentration at higher substrate concentrations.
- This is because collisions between substrate molecules and enzymes ready to accept substrate no longer occur with a likelihood approaching 1. Instead, some collisions occur that would have resulted in the substrate being acted upon if only the enzyme wasn't already occupied.
At even higher substrate concentrations, collisions between enzymes and substrate occur so often that as soon as the enzyme is no longer occupied, a new substrate collides and takes up residence. Consequently, enzyme activity plateaus at very high substrate concentrations at a rate that is independent of absolute substrate concentration.
Enzyme activity once substrate is no longer limiting is instead dependent on what is called the turnover rate of the enzyme, the rate at which substrate cycles through enzyme. See illustration below.
Illustration, saturable enzyme activity
- A large variety of substances act as inhibitors of enzyme activity (i.e., they prevent enzymes from doing their job).
- Some inhibitors occur naturally in cells and are used to control metabolism.
- Some inhibitors are made by other organisms are used to poison the metabolism of susceptible, competing microorganisms (such is the reason that many antibiotics have evolved in various organisms).
- Metabolic wastes can also serve as enzymatic inhibitors, especially those found in environments which are rarely refreshed. See fermentation.
- Some inhibitors act reversibly, some act irreversibly.
- Generally if reversible, an inhibitor may be diluted away and in situ used to fine tune metabolism.
- Continued high or inappropriate concentrations of even reversibly acting inhibitors can cause death. Many antibiotics work this way.
- Some inhibitors bind at the active site. These are called competetive inhibitors.
Feedback inhibition [negative feedback]
- Some inhibitors bind at a site other than the active site. These are called allosteric inhibitors.
The science of control which includes such constructs as negative feedback is called cybernetics.
- In Feedback inhibition (a.k.a., negative feeback) is the Inhibition of enzyme activity in which the products of a reaction or series of reactions acts upon the enzyme(s) responsible for the generation of that product.
- Thus, the more product there is, the less product which is produced. If similarly, the less product there is, the more product which is produced, then there should exist a stable product concentration which is (or range of concentrations which are) maintained over time.
- Feedback inhibition generally leads to well controlled metabolic pathways.
- Your furnace and thermostat at home constute a negative feedback system. The furnace heats things up. At a given temperature the furnace is shut down by the thermostat. The system only starts up again when the inhibitor (the heat) is lost from the system.
- Example: driving at the speed limit:
- An analogy is driving down the highway:
- If you are going too fast, you slow down.
- If you are going too slowly, you speed up.
- Here your velocity is the product, your car is the enzyme (gasoline and air are your substrates), and you are translating your knowledge of your vehicle's velocity into feedback inhibition of the rate at which your car acts upon gasoline and air.
- The product of one or a series of enzymatic reactions acts upon the enzymes responsible for the generation of that product to increase the activity of one or more of these enzymes.
- Positive feedback can lead to out of control situations. Positive feedback tends to be employed by life only under circumstances in which a gross over response (often destructive) is desirable.
- A car analogy would have you accelerating if you were driving too fast.
- Another analogy is the feedback you hear if the microphone and the speaker it is driving are too near each other. The whistle (feedback) that develops is a result of an original noise being picked up by the microphone, amplified, and then directed out of the speakers. The noise coming out of the speakers is also picked up by the microphone upon which it is further amplified, and on and on until the repeatedly amplified noise coming out of the speaker is painfully loud. This analogy isn't quite perfect because here the noise is both the substrate and the product while the amplifier is the enzyme. Note, however, that the speaker cannot become infinitely loud thus implying that the rate of product production is (ultimately) not a linear function of substrate concentration.
- "Positive feedback occurs durign childbirth as the pressure of the infant's head against the exit from the womb stimulates stretch-sensitive receptors. These receptors signal for the release of a hormone from the brain (oxytocin) that intensifies labor contraction. The contractions cause the release of additional hormone and continue until stretching is stopped by the infant's birth." (p. 123, Benjamin et al., 1997)
Facts About Enzymes
Enzymes (computer programs)
Enzymes: Classification, Structure, Mechanisms
Substrates Inhibitors and Enzymatic Related Subjects
Enzyme Biochemistry Chapter
Characteristics of Enzymes
Enzyme Slide Presentation
- Having a uniform composition throughout. Here I use this term to imply that all of the macromolecules in a sample behave identically. This is, of course, is likely a gross over-simplification in most situations, one on par with such things as frictionless planes (physics), completely informed consumers (economics), and published atomic masses that actually describe your sample to the four significant figures (chemistry).
- Active site
- Competetive inhibitor
- Enzyme activity
- Enzyme specificity
- Feedback inhibition
- Negative feedback
- Optimal enzyme activity
- Positive feedback
- Saturable enzyme activity, illustration
Practice question answers
- The specificity of an enzyme is very often a function of (circle correct answer) [PEEK]
- its activity
- its active site's shape
- peptide bond diversity
- the kind of substrate molecules present
- all of the above
- none of the above
- Enzyme X, an isomerase, converts substrate A into product B. Product B binds allosterically to enzyme X strongly and rapidly. Once product B is bound, enzyme X is catalytically active (i.e., can convert A to B). However, if product B is not bound to enzyme X, then enzyme X is not catalytically active (i.e., cannot convert A to B). Given the existence of low concentrations of product B, what two things may determine the activity of any given enzyme X to which a product B has allosterically bound? Be specific. [PEEK]
- In a very general sense, how do enzymes effect catalysis? [PEEK]
- they bring reactants into close proximity
- they tie reactions together
- they stabilize intermediate structures
- they lower activation energy
- all of the above
- none of the above
- Which generally leads to well controlled metabolic pathways? [PEEK]
- substrate concentration
- feedback inhibition
- activation of enzyme by product
- activation of enzyme by substrate
- all of the above
- none of the above
- Which of the following reactions is a coupled reaction? Hint: reactions a and e were not presented in class (i.e., note that they do not involve ATP hydrolysis to ADP + Pi and, unlike the apparently similar reactions presented in class, reactions a and e below are stoichiometrically balanced). [PEEK]
(i) glucose + ATP ----------- glucose-6-phosphate + ADP + heat
(ii) glutamate + NH(4)+ + ATP ------------ glutamine + ADP + Pi + H+ + heat
(iii) Fructose 1,6-diphosphate <======== dihydroxyacetone phosphate +
(iv) glucose-6-phosphate <============== fructose 6-phosphate
(v) fructose 6-phosphate + ATP --------------- Fructose 1,6-diphosphate
+ ADP + heat
- Which of the following reactions or coupled reactions is most likely easily reversed? (circle one correct answer) [PEEK]
- one which generates a great deal of heat.
- one which requires the hydrolysis of ATP.
- one which liberates a great deal of energy.
- one which is significantly exergonic.
- one in which the energy associated with the chemical bonds found in the reactants is similar to that associated with the chemical bonds found in the products.
- one which is significantly endergonic.
- The place in/on an enzyme where substrate specificity is defined and stabilization of reaction intermediates occurs is called the __________? (less than five word answer) [PEEK]
- Competetive inhibitors of enzyme activity general bind to the __________ of enzymes? [PEEK]
- ii, its active site's shape
- concentration of substrate A at lower substrate A concentrations and Enzyme X turnover rate at saturating substrate A concentration.
- v, all of the above
- feedback inhibition
- ii, note that is the only one of those presented which actually consists of two separate reactions: the hydrolysis of ATP plus the synthesis of glutamine from glutamate and ammonium ion plus an input of ATP supplied energy. Note also that I've added the word "heat" to the right hand side of three of the equations as a way of explaining why these equations (but not the other two) are not reversible, i.e., they generated a significant excess of heat that cannot be recaptured as usable energy.
- v, one in which the energy associated with the chemical bonds found in the reactants is similar to that associated with the chemical bonds found in the products.
- active site.
- active site.
- Benjamin, C.L., Garman, G.R., Funston, J.H. (1997).Human Biology. McGraw-Hill Co. Inc., New York. p. 123.
- Raven, P.H., Johnson, G.B. (1995). Biology (updated version). Third Edition. Wm. C. Brown publishers, Dubuque, Iowa. pp. 141-152.
- Tortora, G.J., Funke, B.R., Case, C.L. (1995). Microbiology. An Introduction. Fifth Edition. The Benjamin/Cummings Publishing, Co., Inc., Redwood City, CA, pp. 104-110.