Bovine lung thrombomodulin is purified and used to investigate the basis of the change in substrate specificity of bovine thrombin when bound to thrombomodulin. Bovine thrombomodulin is a single polypeptide having an apparent molecular weight of 84,000 and associates with thrombin with high affinity and rapid equilibrium, to act as a potent cofactor for protein C activation and antagonist of reactions of thrombin with fibrinogen, heparin cofactor 2, and hirudin. Bovine thrombomodulin inhibits the clotting activity of thrombin with Kd less than 2.5 nM. Kinetic analysis of the effect of bovine thrombomodulin on fibrinopeptide A hydrolysis by thrombin indicates competitive inhibition with Kis = 0.5 nM. The active site of thrombin is little perturbed by thrombomodulin, as tosyl-Gly-Pro-Arg-p-nitroanilide hydrolysis and inhibition by antithrombin III are unaffected. Insensitivity of the reaction with antithrombin III is likewise observed with thrombin bound to thrombomodulin on intact endothelium. Antithrombin III-heparin, human heparin cofactor 2, and hirudin inhibit thrombin-thrombomodulin more slowly than thrombin. These effects may arise from a decrease in Ki of the inhibitors for thrombin-thrombomodulin or from changes in the active site not detected by tosyl-Gly-Pro-Arg-p-nitroanilide or antithrombin III. Bovine prothrombin fragment 2 inhibits thrombin clotting activity (Kd less than 7.5 microM) and acts as a competitive inhibitor of protein C activation (Kis = 2.1 microM). The data are consistent with a mechanism whereby thrombomodulin alters thrombin specificity by either binding to or allosterically altering a site on thrombin distinct from the catalytic center required for binding or steric accommodation of fibrinogen, prothrombin fragment 2, heparin cofactor 2, and hirudin.
Fundamentals of Enzyme Kinetics, 4th Edition , Cornish-Bowden, Athel, Wiley-Blackwell, 2012, 510 pp., ISBN 978-3-527-33074-4 (Paperback $79.95). Henry Jakubowski*, * Chemistry Department, College of Saint Benedict/Saint, John's University, Saint Joseph, Minnesota 56374. Enzyme kinetics comprises the major quantitative part of a typical biochemistry course, but as it is usually compartmentalized into a few chapters of a giant tome, it can be tempting for students to either dismiss or not fully appreciate its usefulness in helping investigators decipher and understand biomolecular interactions. It is hard to convey to students, how much kinetic analyses, which require little more than a crude enzyme, a stopwatch, and a collection of substrates and potential inhibitors, can illuminate our understanding of biological processes. What a pity since without an understanding of the kinetic equations and rate constants that characterize simple and complex enzyme-catalyzed reactions, how can students ever hope to really cope with modeling the complexities of biological systems controlled by intertwined signal transduction networks? Any book that provides those who teach or perform enzyme kinetics with reinforcing and strengthening insight and guidance, and that also provides clear and new examples for use in teaching or research is very welcome. Athel Cornish-Bowden's latest edition (4th) of Fundamentals of Enzyme Kinetics does all that and more. In the preface, Cornish-Bowden states that his goal is to provide an understanding of enzyme kinetics, not an encyclopedic coverage of this field. This edition is a complete revision and expansion of earlier versions done in part make it “user friendly” in its style and organization. He succeeds in both tasks. This is a great book for advanced undergraduate students, and for graduate students, faculty, and researchers who truly wish a deeper understanding of this field. From an organizational perspective, the book is a treat. It is extremely readable. Chapter 1 presents a clear review of chemical kinetics for reactions not catalyzed by enzymes, and is followed by Chapter 2 entitled Introduction to Enzyme Kinetics. Noting that a great deal of the theory of enzyme kinetics was developed before a clear indication of the chemical nature of the catalyst, he next presents a brief foray into “alternative” enzymes, including abzymes, ribozymes, and synthetic catalysts. Chapter 4 then deals with practical aspects of kinetics. A very lucidly written explanation of the use of the King–Altman method in deriving steady state rate equations is laid out in Chapter 5, complete with an optional section on the use of determinants in deriving King–Altman patterns. Subsequent chapters cover all the important areas of enzyme kinetics including enzyme inhibition and activation, multi-substrate reactions, pH and isotope effects, and enzyme regulation. He concludes with two important chapters on fast reactions and one that covers “data analysis in an age of kits” which gives a much needed explanation of error analysis. Each chapter ends with bulleted summary points and problems with answers provided at the end of the book. What enhances its readability is Cornish-Bowden's decision to expand the number of figures and tables, and to place many of them, along with references, in the margins of the page corresponding to the written explanations of the figures. The abundance and placement of figures make this book unique and highly effective in leading readers to the understandings he wishes us to achieve. The author has included many graphs consisting of straight line transformations of nonlinear equations, which he argues in his preface allows visible expression of “unseeable” experimental data in ways that computer fitting of data does not easily allow. It is in the understandings provided by the written explanations with support from clear figures that the book shines. He peppers the book with many examples of common misinterpretation and misuses of kinetic data and their transformations. In Chapter 2, he shows how many textbooks misdraw the hyperbolic dependency of initial velocity on substrate concentration, which gives the false impression that saturation of an enzyme is readily achieved and that Vmax is readily determined. He also takes umbrage at the use of Vmax, as the graph of vo versus S reaches a mathematical limit, not a mathematical maximum. Hence, he drops subscripts for velocity terms and uses v and V to represent the initial velocity at any substrate concentration and limiting velocity at saturating values of substrate, respectively, while retaining the “M” in the Michaelis constant, KM. Likewise, he gives an excellent description of the specificity constant, kcat/KM, as the ratio of second order rate constants for competing substrates that are both simultaneously present, a physiologically relevant definition as enzymes do “encounter” this scenario. Also in Chapter 2, the author describes the strengths and liabilities, as well as the historical development, of graphs of v versus A, v versus log A, and linear transformations of the Michaelis–Menten equation. He also has a nice review of progress curve analysis in the presence and absence of product inhibition and explains in clear detail the problems with using this method. For example, he discusses how large errors in V and Km will occur if factors other than product inhibition (such as small losses of enzyme activity or changes in pH or temperature during the measurements occur). It would have been beneficial if he had included a critical review of the recent use of Lambert functions which give product concentration as an explicit function of substrate concentration and allows for determination of kinetic constants in a single time course. The book offers much practical advice. For example in Chapter 4, the author describes straight forward ways to detect changes in enzyme activity during an assay, an occurrence which if unrecognized can wreck the best assays and subsequent analyzes. He notes that investigators cannot assume that site-directed mutants retain their activity during the course of an assay as laboratory selection for an altered activity is quite different from natural selection used in the evolution of wild type enzymes. He gives an excellent explanation of the difficulties in estimating initial rates and the biases in doing so that skew their accurate determination. Nothing is arguable more important to practitioners whose entire analyzes would be nullified by inaccurate determination of initial rates. Chapter 6 deals with reversible inhibition and activation. Noncovalent reversible inhibition is discussed in all undergraduate biochemistry classes. Those who use Cleland's nomenclature for inhibition constants (Kis where s represents a change in the slope of a double reciprocal plot characteristic of competitive inhibition and Kii where i represents the change in the intercept of the plots, found in uncompetitive inhibition) might be pleased to change to the subscripts used by Cornish-Bowden (Kic for competitive inhibition where inhibitor binds to free E and Kiu for uncompetitive where inhibitor binds to enzyme–substrate complex). He goes through a clear treatment of plotting inhibition data, and the relationship between IC50 values and calculated inhibitor dissociation constants for competitive, uncompetitive, and mixed inhibition. As he indicates a heightened need for an understanding of enzyme kinetics in biotechnology and drug design, he devotes part of Chapter 7 to drugs as inhibitors and notes, as he has done in previous papers but which is not discussed in biochemistry textbooks, that inhibition of pathways in vivo is best effected under conditions of constant flux by uncompetitive inhibitors of key central enzymes in the pathway. I could continue with his excellent treatment of multisubstrate reactions and other topics in the second half of the book. I again note the excellent chapter on error analyses at the end of the book. Without knowing the significance of the fit parameters, how can one have confidence in the calculated kinetic parameters or in the explanatory kinetic models? His own errors are on his mind as well as he has provided a list of corrections for his book at http://bipcnrs-mrs.fr/bip10/fek.htm. In addition, he has provided a companion website at http://www.wiley-vch.de/home/fundenzykinet which gives a link to Supporting Information from which all figures and tables can be downloaded for teaching purposes. This book is a gem. The writing is clear and concise. It is filled with historical information, hints, common errors of analysis, and enough theory to enable his readers to do what Cornish-Bowden really wants, which is to understand, perform, and interpret enzyme kinetic analyses correctly and in a way which unleashes the explanatory power derived from their sound use.
The ability to understand, interpret, and create molecular images is a critical learning objective in biochemistry and molecular biology courses. Yet biomolecular visualization skills are rarely explicitly taught. Over a two‐year period, the BioMolViz group organized seven one‐day workshops to explore and establish best practices for the generation, revision, and finalization of biomolecular visualization (BMV) assessments. Our process utilizes a BMV framework (biomolviz.org) to build targeted assessments that address specific learning goals and objectives. We then revise the assessments through a peer review process and integrate a final analysis of images for accessibility to those who are colorblind. Although there is a wealth of information to guide the creation of multiple‐choice questions, we have found that there are few resources for the generation and evaluation of other types of assessments. For example, BMV assessments are more likely to be open‐ended and specifically target molecular visualization skills (e.g. require the student to create an image of DNA or identify a structural feature on a given image of a protein). We invite participants to help us address this gap and refine our method for generating assessments by attending our upcoming three‐day NSF‐funded summer workshops. Activities will include revising existing assessments and creating instruments of interest to new participants. In addition to producing high‐quality BMV assessments aligned with the framework, these workshops will offer a unique professional development for instructors interested in promoting visual literacy by building a community of practice for the instruction of BMV skills. Support or Funding Information This work was supported by the National Science Foundation under award numbers DUE‐1022793, ‐1323414, ‐1503811, IUSE‐1712268, and RCN‐UBE‐1920270. Any opinions, findings, and conclusions or recommendations expressed in this publication are those of the authors and do not necessarily reflect the views of the National Science Foundation.
Abstract Content and emphases in undergraduate biochemistry courses can be readily tailored to accommodate the standards of the department in which they are housed, as well as the backgrounds of the students in the courses. A more challenging issue is how to construct laboratory experiences for a class with both chemistry majors, who usually have little or no experience with biochemical techniques and biology and biochemistry majors who do. This manuscript describes a strategy for differentiating biochemistry labs to meet the needs of students with differing backgrounds. BIOCHEMISTRY AND MOLECULAR BIOLOGY EDUCATION Vol. 39, No. 3, pp. 216–218, 2011.
Climate change caused predominately by carbon dioxide (CO2) from fossil fuel use is a critical issue for our future. It is incumbent on science educators to learn about it and teach it in ways that illustrate the power of science to understand climatic changes and model past, present, and possible climate futures. It is equally important for educators to address alternative explanations that do not cause present warming. We provide sufficient background to understand the effects of atmospheric CO2 on climate, how we know past values of both CO2 and temperature, and how mathematical models lead to a quantitative understanding and predictions of increases in temperature on doubling atmospheric CO2. We discuss alternative but incorrect explanations for present warming that are employed by those who use misinformation and disinformation to take attention away from the main cause, the burning of fossil fuels. Guided student activities are provided to help students develop an understanding of the causes and strategies to mitigate the worst effects of climate change. We must provide students with a sound understanding of climate change to empower them with the knowledge to make effective choices. Informed students can develop into agents of change as we prepare them to live in their changing climate futures.
ADVERTISEMENT RETURN TO ISSUEPREVArticleNEXTThe Study of Lipid Aggregates in Aqueous Solution: Formation and Properties of Liposomes with an Encapsulated Metallochromic DyeHenry V. Jakubowski , Mary Penas , and Kenneth Saunders Cite this: J. Chem. Educ. 1994, 71, 4, 347Publication Date (Print):April 1, 1994Publication History Received3 August 2009Published online1 April 1994Published inissue 1 April 1994https://doi.org/10.1021/ed071p347Request reuse permissions Article Views662Altmetric-Citations9LEARN ABOUT THESE METRICSArticle Views are the COUNTER-compliant sum of full text article downloads since November 2008 (both PDF and HTML) across all institutions and individuals. These metrics are regularly updated to reflect usage leading up to the last few days.Citations are the number of other articles citing this article, calculated by Crossref and updated daily. Find more information about Crossref citation counts.The Altmetric Attention Score is a quantitative measure of the attention that a research article has received online. Clicking on the donut icon will load a page at altmetric.com with additional details about the score and the social media presence for the given article. Find more information on the Altmetric Attention Score and how the score is calculated. Share Add toView InAdd Full Text with ReferenceAdd Description ExportRISCitationCitation and abstractCitation and referencesMore Options Share onFacebookTwitterWechatLinked InReddit PDF (3 MB) Get e-Alertsclose Get e-Alerts
While molecular visualization has been recognized as a threshold concept in biology education, the explicit assessment of students' visual literacy skills is rare. To facilitate the evaluation of this fundamental ability, a series of NSF-IUSE-sponsored workshops brought together a community of faculty engaged in creating instruments to assess students' biomolecular visualization skills. These efforts expanded our earlier work in which we created a rubric describing overarching themes, learning goals, and learning objectives that address student progress toward biomolecular visual literacy. Here, the BioMolViz Steering Committee (BioMolViz.org) documents the results of those workshops and uses social network analysis to examine the growth of a community of practice. We also share many of the lessons we learned as our workshops evolved, as they may be instructive to other members of the scientific community as they organize workshops of their own.
