7 Structural Changes and Metabolism of Proteins Following Heat Denaturation J. N. NEUCERE and J. P. CHERRY
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United States Department of Agriculture, Southern Regional Research Center, Agricultural Research Service, New Orleans, LA 70179
Heat is perhaps the primary factor that enhances the safety, nutritional quality, and palatability of most foods. Mild or moderate temperatures can be beneficial in achieving these goals, whereas, extreme temperatures often are deleterious in terms of biological value and toxicity. The complex reactions that occur during extreme heat treatments, which usually involve simple and intricate molecules and ions, modify the physicochemical properties and metabolic efficiency of proteins. A key concept in protein chemistry is the effect heat has on active centers or sites within these molecules. These centers, involving any number of functional groups, are defined as the regions of space where the binding of other molecules and ions occur. Some of the most reactive sites in proteins are amine, sulfhydryl, tyrosyl, and imidizole groups. Heat-induced changes in any or all of these functional groups are directly related to physicochemical modifications such as solubility, enzymatic activity, antigenic reactivity, electrophoretic migration and association-dissociation constraints. Anti-nutritional parameters and toxicological aspects of products from protein heated in the presence of carbohydrates are well recognized from the many studies via the Amadori and Maillard reactions. The formation of diverse isopeptides induced by heat are also critical factors that can adversely affect the biological value of proteins. Measuring toxicity or harmful effects, for example, of such complicated reaction products is not a simple matter. Adversity could involve either a modified intact protein or the presence of intricate amino acids and peptides that have formed. This chapter concentrates on several empirical views of structural changes and anti-nutritional aspects of proteins under various conditions of heat treatments. Experimental Procedures The reader is referred to the fol 1 owing publications on techniques of gel electrophoresis Q), immunochemistry (j2), Current address: United States Department of Agriculture, Eastern Regional Research Center, Philadelphia, PA 19118. 1
This chapter not subject to U.S. copyright. Published 1982 American Chemical Society.
In Food Protein Deterioration; Cherry, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1982.
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functionality (3) and nutrition (4). Studies on the determination of multiple forms of enzymes and t h e i r use as biological indicators for food uses and applications were reviewed by Cherry ( ! , £ ) . Whole seeds, f u l l - f a t and fat-free meal s, concentrates and/or isolates are used in research to determine the effects of moist and dry heat on protein properties. For example, peanut kernels are moist-heated in water in a temperature-control 1ed steam retort kept at various temperatures for different time intervals (7). After moist heat treatment, the seals are lyophilized and then ground into a meal for protein analyses. Dry heat treatments may be conducted by placing seeds in a forced-draft oven at various temperatures and time intervals ( £ ) ; purified proteins such as peanut α-arachin are s i m i l a r l y heat-treated (9). Seeds can be made to imbibe varying amounts of water to 40% and then dry heated (8). Microwave heating of seeds containing only t h e i r innate moisture or after they have been placed in water is conducted at operational frequencies as high as 2,480 MHz (10). Evidence of Protein Structural Changes Induced by Heat S o l u b i l i t y . Perhaps the most obvious evidence of conforma tional changes i n protein structure are the alterations in protein s o l u b i l i t y . In general, protein s o l u b i l i t y decreases with time and temperature of heat treatment; and considerable variations i n the degree of change exist among proteins. Interactions with several functional groups ( e . g . , SH- or tyrosyl-residues) become more prevalent fol 1 owing unfolding of protein chains, resulting in diverse and complicated precipitation reactions. The exact mechanisms leading to precipitation are most d i f f i c u l t to study after heat treatments because the process is usually i r r e v e r s i b l e . It is well known that the interactions of water molecules with ionic and polar groups of proteins strongly inf1uence the folded conformations of proteins (11). Consequently, moist heat usually has a more complex effect on the s o l u b i l i t y properties of pro teins than does dry heat, and is strongly influenced by ionic strength and pH. An example of s o l u b i l i t y changes of peanut proteins induced by heat is shown in Figure 1. In this study (8), seeds were al 1 owed to imbibe d i s t i l l e d water for 16 hr at 25°C to a final moisture content of 40% and were heated for 1 hr from 110 to 155°C. Other seed samples were heated on an "as is" basis (5% moisture) under identical conditions. S o l u b i l i t y of total pro tein decreased almost 1inearly with temperature for the dry-heated seeds. For the wet-heated samples, the relationship was quite d i f f e r e n t ; a plot of s o l u b i l i t y versus temperature showed a sigmoid-1ike curve with a minimum at 120°C decreasing sharply after 145°C. Compared to the 110°C wet- and dry-heated samples, the control had an intermediate degree of s o l u b i l i t y . This information implies that the i n i t i a l steps of denaturation i n the
In Food Protein Deterioration; Cherry, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1982.
Downloaded by NORTH CAROLINA STATE UNIV on May 3, 2015 | http://pubs.acs.org Publication Date: December 13, 1982 | doi: 10.1021/bk-1982-0206.ch007
7.
NEUCERE AND CHERRY
Heat Denaturation
of
Proteins
140
150
137
40.0H
35.0
g 30.0h
— 25.0 Ο Ζ Ο Ο 20.0h Ζ UJ
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15.0
01 CL 10.0
5.0
UN
IÏÔ
120
130
160
TEMP. °C Figure 1. Relative protein solubilities of intact peanuts, after moist (O) and dry (Φ) heat. Triangles indicate native seed or control. (Reproduced, from Ref. 9. Copyright 1974, American Chemical Society.)
In Food Protein Deterioration; Cherry, J.; ACS Symposium Series; American Chemical Society: Washington, DC, 1982.
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imbibed seed involve a gradual change of the water-protein system within the seed between 110 and 120°C. Beyond this point, a more complex water-protein system appears to develop possibly involving protein fragments and other macromolecules. At low moisture con tent, a continuous change in the water-protein system seems to occur. Cherry et
