In the Classroom
Using Denatured Egg White as a Macroscopic Model for Teaching Protein Structure and Introducing Protein Synthesis for High School Students Paulo R. M. Correia* Escola de Artes, Ciências e Humanidades, Universidade de São Paulo, 03828-080, São Paulo, SP, Brazil; *
[email protected] Bayardo B. Torres Instituto de Química, Universidade de São Paulo, 05513-970, São Paulo, SP, Brazil
The denaturation of proteins is an important biochemical process that involves changes in the native shape of the molecule and the loss of biological activity. The interaction between amino acids is responsible for the native three-dimensional structure of a protein, which can be substantially altered by changes in the environment even if its amino acid sequence remains unchanged (17). Introducing this phenomenon to high school students is difficult owing to the lack of appropriate macroscopic models for mediating the discussion of protein molecular structure, which requires a high level of abstraction. In this article we describe a set of classroom activities that were devised using low-level abstraction media to introduce protein structure and denaturation for high school students. During three 50-minute classes, the teacher performs two experimental demonstrations, the students assemble a macroscopic model for studying protein structure, and they perform a role-play to represent protein synthesis in a cell. All of these activities have a low degree of abstraction and were selected based on Dale’s cone of experience. The students should have some background knowledge about chemical bonding (covalent and ionic), intraand intermolecular forces (mainly hydrogen bonds), and organic
older
verbal symbols
high
visual symbols
still pictures motion picture: edited, mediated reality educational television: real-time, mediated reality exhibits: edited reality study trips, viewing reality demonstrations: learner becomes a spectator
Degree of Sophistication
radio and recordings
Age
One challenging aspect of teaching chemistry for high school students is to explain chemical phenomena including the behavior of atoms and molecules (1, 2). Overcoming this pivotal difficulty is critical since the comprehension of atomic and molecular events, such as the interaction of reagents during a chemical reaction, the chemical bonding of atoms, and intermolecular forces is essential to achieving a full understanding of matter from a chemical point of view. The successful accomplishment of teaching molecular and atomic facts depends on the didactical strategy and the media adopted in consideration of the level of abstraction of the subject to be taught and the students’ capability to deal with abstract operations. Dale’s cone of experience (Figure 1) can be useful as a reference when planning classroom activities (3) and for establishing adequate communication in the classroom during the learning process. Because most upper secondary students are not completely prepared to make abstract operations involving microscopic entities, the lack of concrete activities during a chemical discussion at the molecular level can impair their understanding. Thus, interesting but complex chemical events are usually not fully explored in high school. Nevertheless, the choice of concrete experiences can support the learning process in this situation (2, 3). Biochemistry has been extensively discussed by the mass media and public interest in this subject has also increased. The majority of publications dealing with biochemistry teaching are devoted to undergraduate students (4–10). However, interdisciplinary topics involving this field, which merges chemistry and biology, can be explored in high school as well (11–15) enhancing the students’ motivation by linking scholarly knowledge to everyday facts (1). Moreover the perception of chemistry as a basis for many current biological scientific developments gives more meaning to the chemical subjects discussed in school (16). Among all topics related to the recent scientific advances of molecular biology (the human genome project, stem cells) and recombinant DNA technology (cloning, transgenic organisms, gene therapy), proteins have gained prominence among the general public (17–23). Indeed, proteins are the sole expression of a gene. Additionally, in the past few years as awareness of physical well-being has increased, greater attention is being paid to nutritional habits and proteins are again prominent in consideration of their essential amino acid content. The understanding of proteins as molecules, and their importance for living organisms from a biochemical point of view, can lead high school students to critically evaluate the importance of the scientific research on this topic.
dramatized experiences: engaging, qualitative
younger
contrived experiences: representation of reality direct purposeful experiences multi-sensory, highly qualitative
low
Figure 1. Dale’s cone of experience and its correlation with the students’ age and the abstraction level.
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In the Classroom
Procedures
functional groups (amines and carboxylic acids) to make more the proposed activities more meaningful.
Class 1: Experimental Demonstrations The connection between students’ background comprehension and the chemical nature of proteins was made by two demonstrations involving chicken egg albumin that were performed by the teacher. These demonstrations were the basis for all planned activities. After separating the egg white from the yolk, the denaturation of albumin was observed by (i) increasing the temperature and (ii) changing the original solvent (water) to ethanol. In both cases, the appearance of a white rubbery product was observed. The students were asked to explain the change in appearance of albumin after heating and after changing the solvent.
Assessing the Students’ Background Before starting the activities, the students were asked to answer some questions to assess their understanding about proteins. The evaluation of 55 feedback forms showed a disconnect between the “protein” presented during biology classes (as a cellular structure) and the “protein” discussed during chemistry classes (as a molecule). In spite of the common topic, the students cannot merge these different approaches. Possible reasons are a lack of interdisciplinary activities and the presentation of this subject in separate courses at different times. The biological approach dominates the students’ view of proteins perhaps because it does not explore the molecular nature of proteins, requiring only low-abstraction cognitive operations. As a consequence, the digestive enzymes appeared as the most important (if not the unique) example of proteins in living organisms. Moreover, since proteins were not considered as molecules, students showed a poor understanding of protein denaturation. On the other hand, the chemical approach requires a higher level of abstraction and the use of microscopic entities. Considering the students’ capability to deal with high-level abstract operations, and the requirements to discuss proteins at the molecular level, the traditional didactical tools should be replaced by more concrete activities to assist the learning process. Merging the chemical point of view to the established biological concept of protein is only possible when the instructional activities are planned using concrete experiences to broaden the students’ background knowledge of the molecular nature of proteins.
Guided Discussion Following the demonstrations, a guided discussion with all students identified the observed visual changes indicating the occurrence of a chemical transformation. Thus, the constituents of the egg white can be considered as molecules that can be transformed by heating or by changing the solvent. Proteins were presented as amino acid polymers that are linked by peptide bonds established between the carboxyl group of one amino acid and the amino group of the other. Class 2: Enriching the Molecular Understanding of Protein Denaturation A macroscopic model for a 10 amino acid peptide was built from colored paper clips and modeling clay. The students were divided into small groups to favor collaborative learning (24–26). Each group received a specific amino acid sequence (Table 1) to be assembled using colored paper clips according to the information in Table 2.
Table 1. Amino Acid Sequences Assembled by the Students Peptide Seq Seq Seq Seq Seq Seq
1 2 3 4 5 6
#1
#2
#3
#4
Thra Ala Gly Ala Gly Thra
Cys Sera Ala Cys Lys Cys
Ala Gly Ala Thra Ala Ala
Cys Cys Cysb Cys Ala Cys
aHydrogen-bond
formation is possible with these amino acids
Amino Acid #5 #6 Gly Gly Cysb Gly Gly Gly bActive
Cys Thra Asp Gly Cysb Cys
#7
#8
#9
#10
Ala Cysb Ala Ala Cysb Ala
Ala Cysb Cys Sera Asp Ala
Thra Ala Lys Gly Gly Thra
Gly Gly Gly Ala Ala Gly
site in the peptide; amino acids highlighted with bolded font.
Table 2. Correspondence between the Amino Acid and the Paper Clip Color Paper Clip Color
Amino Acid
Characteristic
Chemical Structure
Blue
Cysteine (Cys)
Polar
HSCH2–CH(NH3+)CO2‒
Green
Serine (Ser)
Polar
HOCH2–CH(NH3+)CO2‒
Yellow
Threonine (Thr)
Polar
CH3CH(OH)–CH(NH3+)CO2‒
Red
Alanine (Ala)
Nonpolar
White
Aspartic acid (Asp)
Ionic
Black
Glycine (Gly)
Nonpolar
Orange
Lysine (Lys)
Ionic
CH3–CH(NH3+)CO2‒ HO2CCH2–CH(NH3+)CO2‒ H2–CH(NH3+)CO2‒ NH2CH2CH2CH2CH2–CH(NH3+)CO2‒
1942 Journal of Chemical Education • Vol. 84 No. 12 December 2007 • www.JCE.DivCHED.org
In the Classroom
The primary structure of proteins was presented as the linear sequence of all amino acids (paper clips in Figure 2, top). Then the students were informed about the amino acid structures and the possibility of interaction among these amino acids when, for example, hydroxyl groups are present in the amino acid side chains that allow hydrogen-bond formation. This interaction between different amino acids was represented by joining the paper clips together using modeling clay (Figure 2, bottom). Other possibilities of interaction among the amino acids were also explored by considering the molecular structures of the amino group (−NH3+) and carbonyls (Tables 1 and 2). Finally, the existence of an active site was also declared and it was supposed to be formed by two consecutive cysteines linked in a sequence. Therefore, students were able to see the primary and the three-dimensional protein structures, as well as identify the active site as a part of the molecule. The denaturation process was macroscopically represented using the model by removing the modeling clay and changing the spatial structure of the amino acid sequence (Figure 2). Guided Discussion The proteins in egg white (mainly albumin) are globular proteins, which means that the long protein molecule is twisted, folded, and curled up into an approximate spherical shape (17). A variety of weak chemical bonds, mainly hydrophobic interactions, keep the protein tightly curled as it drifts placidly in the water that surrounds it. When heat is applied, those placidly drifting egg white proteins are agitated, which bounces them around. They slam into the surrounding water molecules and they bash into each other. All of this bashing about breaks the weak bonds that maintain the protein’s native structure. The egg proteins uncurl and bump into other proteins that have also uncurled. New chemical bonds form, but rather than binding the protein to itself, these bonds connect one protein to another. After enough of this bashing and bonding, the solitary egg proteins are no longer solitary. They have formed a network of interconnected proteins. The water in which the proteins once floated is captured and held in the protein web. If the egg is left at a high temperature for too long, too many intermolecular bonds form and the egg white becomes rubbery (27). A similar process occurs when ethanol is added to the egg white without any heating. In this case, the ethanol (much less polar than water) interacts with the hydrophobic side chains of amino acids that are normally interacting with each other deep in the hydrophobic core of the proteins’ native structure. Their newly formed interactions with ethanol trigger a variety of sequential structural disruptions and the formation of different weak chemical bonds that changes the overall structure of the protein and leads to its final aspect similar to the heated egg white. Discussion reinforcing the importance of a protein’s shape and biological catalysis was also presented. The lock and key theory was presented to explain the specific action of an enzyme on a single substrate. Secondary structure was discussed as a regular part of the three-dimensional arrangement of amino acids. The whole structure was also discussed by using the figures from a biochemistry book (17). At this point, the role of proteins as biological catalysts was recalled and the lock and key theory was presented from the molecular point of view. This theory, first postulated in 1894 by Emil Fischer, had already been presented during biology classes, but the students’ understanding was restricted to the macroscopic analogy: the enzyme (protein)
Figure 2. Macroscopic model for a 10-amino acid sequence (top) without interaction and (bottom) with interaction between two amino acid molecules. (Note that the colors suggested in Table 2 do not apply to this figure.)
is the lock and the substrate is the key (17). Only the correctly sized key fits into the key hole (active site) of the lock (enzyme). The concept of the active site was visualized using the macroscopic model, and the students were asked to search for it (two cysteines in a row) within their 10 amino acid sequences. At the end, the students understood the importance of a protein’s molecular shape to preserve the form of the active site. Changes in the interactions among the amino acids can affect the native shape of a protein, and it can lose its biological functionality (as a catalyst, for example). Therefore, the denaturation process was reconstructed with the students from a molecular point of view where it was understood that changes in the three dimensional structure were due to changes in amino acid interactions and not peptide bond cleavage. It is important to stress that this was the first time the students had made this important distinction. Class 3: A Role-Play To Develop the Systemic View of a Cell After emphasizing the importance of protein structure for the proper functioning of living organisms, an activity was done to join together several concepts from molecular biology into a single framework. Hence, the students performed a roleplay (28) involving the synthesis of a protein. Each student represented a specific cellular component and glucagon was synthesized from the teamwork developed by the students. Each one received an ID card corresponding to a cellular structure involved in this biological process: DNA, mRNA, ribosome, and aminoacyl-tRNAs. The glucagon synthesis was carried out by following the sequence: (i) DNA transcription into an mRNA chain in the nucleus, (ii) mRNA travels to the cytoplasm, (iii) the ribosome attaches to the mRNA, (iv) the aminoacyl-tRNAs
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In the Classroom
come to the ribosome in the right order, and (v) the amino acids are linked by peptide bonds. The coding scheme of amino acids using three letters (bases) was explained, and initiation and termination codons were presented before the role-play.
the Conselho Nacional de Desenvolvimento Científico e Tecnológico for the provided fellowship (CNPq 150325∙2004-5) and for the financial support (CNPq 553710∙2006-0).
Guided Discussion The essential amino acids were discussed highlighting the need for ingesting them through a balanced diet to ensure that all of the components required to synthesize a protein will be present in the cells. Additionally, the importance of genomic studies was presented in a broad perspective and relationships between DNA and proteins were clarified. Thus, terms such as genome and proteome became more meaningful for the students after the role-play. Problems related to changes in DNA base sequences were also pointed out, and the modification in a protein’s structure was considered as a harmful consequence. Glucagon was selected based on its size (29 amino acids) and its role as a hormone to expand the protein usefulness beyond the enzymatic purposes that are extensively taught during high school. The importance of controlling glucose concentration in the blood was discussed and the combined action of insulin and glucagon was presented. The molecular regulation of living organisms became more evident, and biochemistry could be truly understood as a merger of chemistry and biology.
Literature Cited
Concluding Remarks The use of macroscopic models is a powerful didactical strategy to represent molecular and atomic events. They can convert microscopic entities into tangible objects, reducing the abstraction level required to discuss chemistry with high school students. Thus, interesting discussions involving molecules and their behavior can take place efficiently when mediated by concrete experiences. The choice of these activities exploited their low degree of abstraction, which can support the learning process in a student-centered approach. The misconceptions observed as background knowledge were changed and the new scientific understanding is in accordance with the acceptable scientific theories used to explain protein denaturation. Moreover, the systemic understanding of a cell allowed the students to integrate disjointed knowledge, and after the role-play, some molecular biology concepts were arranged into a single framework. From gene to protein structure, the students were able to organize the events and molecules involved with protein synthesis, as well as understand the relevance and consequences of the recent advances of genetic research. Acknowledgments The authors are in debt to Colégio Integrado Objetivo from Suzano∙SP; Melissa Dazzani, the teacher responsible for applying all of the activities; and her students. PRMC also thanks
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