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- Volume 41, 2012
Annual Review of Biophysics - Volume 41, 2012
Volume 41, 2012
- Preface
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How Should We Think About the Ribosome?
Vol. 41 (2012), pp. 1–19More LessIn a few years we are likely to have structures for the ribosome in all the conformations it assumes during protein synthesis. The golden age of ribosome structure determination is thus drawing to a close, and as it does the focus in the field will shift from structure determination to understanding why the ribosome's structure changes the way it does as it performs its function. Thus in the future, kinetic and thermodynamic experiments will become increasingly important, and as they do, the field will have to start thinking about the dynamics of the ribosome far more carefully than it has in the past. The reasoning that underlies these assertions will be explained, and a more general issue explored, namely what can be said today about the modus operandi of the ribosome. What kind of a device is it?
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Mechanisms of Sec61/SecY-Mediated Protein Translocation Across Membranes
Vol. 41 (2012), pp. 21–40More LessThe Sec61 or SecY channel, a universally conserved protein-conducting channel, translocates proteins across and integrates proteins into the eukaryotic endoplasmic reticulum (ER) membrane and the prokaryotic plasma membrane. Depending on channel-binding partners, polypeptides are moved by different mechanisms. In cotranslational translocation, the ribosome feeds the polypeptide chain directly into the channel. In posttranslational translocation, a ratcheting mechanism is used by the ER-lumenal chaperone BiP in eukaryotes, and a pushing mechanism is utilized by the SecA ATPase in bacteria. In prokaryotes, posttranslational translocation is facilitated through the function of the SecD/F protein. Recent structural and biochemical data show how the channel opens during translocation, translocates soluble proteins, releases hydrophobic segments of membrane proteins into the lipid phase, and maintains the barrier for small molecules.
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Racemic Protein Crystallography
Vol. 41 (2012), pp. 41–61More LessAlthough natural proteins are chiral and are all of one “handedness,” their mirror image forms can be prepared by chemical synthesis. This opens up new opportunities for protein crystallography. A racemic mixture of the enantiomeric forms of a protein molecule can crystallize in ways that natural proteins cannot. Recent experimental data support a theoretical prediction that this should make racemic protein mixtures highly amenable to crystallization. Crystals obtained from racemic mixtures also offer advantages in structure determination strategies. The relevance of these potential advantages is heightened by advances in synthetic methods, which are extending the size limit for proteins that can be prepared by chemical synthesis. Recent ideas and results in the area of racemic protein crystallography are reviewed.
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Disulfide Bonding in Protein Biophysics
Vol. 41 (2012), pp. 63–79More LessIt has been known for many decades that cell surface, soluble-secreted, and extracellular matrix proteins are generally rich in disulfide bonds, but only more recently has the functional diversity of disulfide bonding in extracellular proteins been appreciated. In addition to the classic mechanisms by which disulfide bonds enhance protein thermodynamic stability, disulfides in certain configurations contribute particular mechanical properties to proteins that sense and respond to tensile forces. Disulfides may help warp protein folds for the evolution of new functions, or they may fasten aggregation-prone flaps of polypeptide to protein surfaces to prevent fibrilization or oligomerization. Disulfides can also be used to package and secure macromolecular cargo for intercellular transport. A series of case studies illustrating diverse biophysical roles of disulfide bonding are reviewed, with a focus on proteins functioning in the extracellular environment.
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Prokaryotic Diacylglycerol Kinase and Undecaprenol Kinase
Vol. 41 (2012), pp. 81–101More LessProkaryotic diacylglycerol kinase (DAGK) and undecaprenol kinase (UDPK) are the lone members of a family of multispan membrane enzymes that are very small, lack relationships to any other family of proteins—including water soluble kinases—and exhibit an unusual structure and active site architecture. Escherichia coli DAGK plays an important role in recycling diacylglycerol produced as a by-product of biosynthesis of molecules located in the periplasmic space. UDPK seems to play an analogous role in gram-positive bacteria, where its importance is evident because UDPK is essential for biofilm formation by the oral pathogen Streptococcus mutans. DAGK has also long served as a model system for studies of membrane protein biocatalysis, folding, stability, and structure. This review explores our current understanding of the microbial physiology, enzymology, structural biology, and folding of the prokaryotic DAGK family, which is based on over 40 years of studies.
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Allostery and the Monod-Wyman-Changeux Model After 50 Years
Vol. 41 (2012), pp. 103–133More LessThe Monod-Wyman-Changeux (MWC) model was conceived in 1965 to account for the signal transduction and cooperative properties of bacterial regulatory enzymes and hemoglobin. It was soon extended to pharmacological receptors for neurotransmitters and other macromolecular entities involved in intracellular and intercellular communications. Five decades later, the two main hypotheses of the model are reexamined on the basis of a variety of regulatory proteins with known X-ray structures: (a) Regulatory proteins possess an oligomeric structure with symmetry properties, and (b) the allosteric interactions between topographically distinct sites are mediated by a conformational transition established between a few preestablished states with conservation of symmetry and ligand-directed conformational selection. Several well-documented examples are adequately represented by the MWC model, yet a few possible exceptions are noted. New questions are raised concerning the dynamics of the allosteric transitions and more complex supramolecular ensembles.
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Protein Structure in Membrane Domains
Vol. 41 (2012), pp. 135–155More LessOf great interest to the academic and pharmaceutical research communities, helical transmembrane proteins are characterized by their ability to dissolve and fold in lipid bilayers—properties conferred by polypeptide spans termed transmembrane domains (TMDs). The apolar nature of TMDs necessitates the use of membrane-mimetic solvents for many structure and folding studies. This review examines the relationship between TMD structure and solvent environment, focusing on principles elucidated largely in membrane-mimetic environments with single-TMD protein and peptide models. Following a brief description of TMD sequence and conformational characteristics gleaned from the structural database, we present an overview of the conceptual models used to study folding in vitro. The impact of sequence and solvent context on the incorporation of TMDs into membranes, and its role in measurements of TMD self-assembly strengths, is then described. We conclude with a discussion of the nonspecific effects of membrane components on TMD stability.
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Bacterial Mechanosensitive Channels—MscS: Evolution's Solution to Creating Sensitivity in Function
Vol. 41 (2012), pp. 157–177More LessThe discovery of mechanosensing channels has changed our understanding of bacterial physiology. The mechanosensitive channel of small conductance (MscS) is perhaps the most intensively studied of these channels. MscS has at least two states: closed, which does not allow solutes to exit the cytoplasm, and open, which allows rapid efflux of solvent and solutes. The ability to appropriately open or close the channel (gating) is critical to bacterial survival. We briefly review the science that led to the isolation and identification of MscS. We concentrate on the structure-function relationship of the channel, in particular the structural and biochemical approaches to understanding channel gating. We highlight the troubling discrepancies between the various models developed to understand MscS gating.
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Cooperativity in Cellular Biochemical Processes: Noise-Enhanced Sensitivity, Fluctuating Enzyme, Bistability with Nonlinear Feedback, and Other Mechanisms for Sigmoidal Responses
Vol. 41 (2012), pp. 179–204More LessCooperativity in classical biophysics originates from molecular interactions; nonlinear feedbacks in biochemical networks regulate dynamics inside cells. Using stochastic reaction kinetic theory, we discuss cooperative transitions in cellular biochemical processes at both the macromolecular and the cellular levels. We show that fluctuation-enhanced sensitivity (stochastic focusing) shares an essential feature with the transition in a bistable system. The same theory explains zeroth-order ultrasensitivity with temporal cooperativity. Dynamic cooperativity in fluctuating enzyme (i.e., dynamic disorder), stochastic focusing, and the recently proposed stochastic binary decision all have a shared mechanism: They are generalizations of the hyperbolic response of Michaelis-Menten kinetics x/(K+x), with fluctuating K or stochastic x. Sigmoidal dependence on substrate concentration necessarily yields affinity amplification for competing ligands; both sigmoidal response and affinity amplification exhibit a square law. We suggest two important characteristics in a noise: its multimodal distribution structure and its temporal irreversibility. The former gives rise to self-organized complexity, and the latter contains useful, albeit hidden, free energy that can be utilized for biological functions. There could be structures and energy in biochemical fluctuations.
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Network-Based Models as Tools Hinting at Nonevident Protein Functionality
Vol. 41 (2012), pp. 205–225More LessNetwork-based models of proteins are popular tools employed to determine dynamic features related to the folded structure. They encompass all topological and geometric computational approaches idealizing proteins as directly interacting nodes. Topology makes use of neighborhood information of residues, and geometry includes relative placement of neighbors. Coarse-grained approaches efficiently predict alternative conformations because of inherent collectivity in the protein structure. Such collectivity is moderated by topological characteristics that also tune neighborhood structure: That rich residues have richer neighbors secures robustness toward random loss of interactions/nodes due to environmental fluctuations/mutations. Geometry conveys the additional information of force balance to network models, establishing the local shape of the energy landscape. Here, residue and/or bond perturbations are critically evaluated to suggest new experiments, as network-based computational techniques prove useful in capturing domain movements and conformational shifts resulting from environmental alterations. Evolutionarily conserved residues are optimally connected, defining a subnetwork that may be utilized for further coarsening.
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Filamins in Mechanosensing and Signaling
Vol. 41 (2012), pp. 227–246More LessFilamins are essential, evolutionarily conserved, modular, multidomain, actin-binding proteins that organize the actin cytoskeleton and maintain extracellular matrix connections by anchoring actin filaments to transmembrane receptors. By cross-linking and anchoring actin filaments, filamins stabilize the plasma membrane, provide cellular cortical rigidity, and contribute to the mechanical stability of the plasma membrane and the cell cortex. In addition to binding actin, filamins interact with more than 90 other binding partners including intracellular signaling molecules, receptors, ion channels, transcription factors, and cytoskeletal and adhesion proteins. Thus, filamins scaffold a wide range of signaling pathways and are implicated in the regulation of a diverse array of cellular functions including motility, maintenance of cell shape, and differentiation. Here, we review emerging structural and functional evidence that filamins are mechanosensors and/or mechanotransducers playing essential roles in helping cells detect and respond to physical forces in their local environment.
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ATP Utilization and RNA Conformational Rearrangement by DEAD-Box Proteins
Vol. 41 (2012), pp. 247–267More LessRNA helicase enzymes catalyze the in vivo folding and conformational re-arrangement of RNA. DEAD-box proteins (DBPs) make up the largest family of RNA helicases and are found across all phyla. DBPs are molecular motor proteins that utilize chemical energy in cycles of ATP binding, hydrolysis, and product release to perform mechanical work resulting in reorganization of cellular RNAs. DBPs contain a highly conserved motor domain helicase core. Auxiliary domains, enzymatic adaptations, and regulatory partner proteins contribute to the diversity of DBP function throughout RNA metabolism. In this review we focus on the current understanding of the DBP ATP utilization mechanism in rearranging and unwinding RNA structures. We discuss DBP structural properties, kinetic pathways, and thermodynamic features of nucleotide-dependent interactions with RNA. We highlight recent advances in the DBP field derived from biochemical and molecular biophysical investigations aimed at developing a quantitative mechanistic understanding of DBP molecular motor function.
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Zero-Mode Waveguides for Single-Molecule Analysis
Vol. 41 (2012), pp. 269–293More LessWe review the optical properties, fabrication, and applications of zero-mode waveguides (ZMWs) for single-molecule studies. These simple nano-structures allow individual molecules to be isolated for optical analysis at high concentrations. Fluorescent species are observed in a sufficiently small volume that the average number of fluorescent molecules is less than one, even at concentrations high enough for biochemical reactions to proceed at normal rates. Arrays of such structures can also be engineered into systems for real-time analysis of large numbers of single-molecule reactions or binding events. We also review the integration of ZMWs in different optical configurations and microfluidic systems.
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Single-Molecule Views of Protein Movement on Single-Stranded DNA
Vol. 41 (2012), pp. 295–319More LessThe advent of new technologies allowing the study of single biological molecules continues to have a major impact on studies of interacting systems as well as enzyme reactions. These approaches (fluorescence, optical, and magnetic tweezers), in combination with ensemble methods, have been particularly useful for mechanistic studies of protein–nucleic acid interactions and enzymes that function on nucleic acids. We review progress in the use of single-molecule methods to observe and perturb the activities of proteins and enzymes that function on flexible single-stranded DNA. These include single-stranded DNA binding proteins, recombinases (RecA/Rad51), and helicases/translocases that operate as motor proteins and play central roles in genome maintenance. We emphasize methods that have been used to detect and study the movement of these proteins (both ATP-dependent directional and random movement) along the single-stranded DNA and the mechanistic and functional information that can result from detailed analysis of such movement.
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Extending Microscopic Resolution with Single-Molecule Imaging and Active Control
Vol. 41 (2012), pp. 321–342More LessSuperresolution imaging of biological structures provides information beyond the optical diffraction limit. One class of superresolution techniques uses the power of single fluorescent molecules as nanoscale emitters of light combined with emission control, variously described by the acronyms PALM/FPALM/STORM and many others. Even though the acronyms differ and refer mainly to different active-control mechanisms, the underlying fundamental principles behind these “pointillist” superresolution imaging techniques are the same. Circumventing the diffraction limit requires two key steps. The first step (superlocalization) is the detection and localization of spatially separated single molecules. The second step actively controls the emitting molecules to ensure a very low concentration of single emitters such that they do not overlap in any one imaging frame. The final image is reconstructed from time-sequential imaging and superlocalization of the single emitting labels decorating the structure of interest. The statistical, imaging, and active-control strategies for achieving superresolution imaging with single molecules are reviewed.
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Metabolite Recognition Principles and Molecular Mechanisms Underlying Riboswitch Function
Vol. 41 (2012), pp. 343–370More LessRiboswitches are mRNA elements capable of modulating gene expression in response to specific binding by cellular metabolites. Riboswitches exert their function through the interplay of alternative ligand-free and ligand-bound conformations of the metabolite-sensing domain, which in turn modulate the formation of adjacent gene expression controlling elements. X-ray crystallography and NMR spectroscopy have determined three-dimensional structures of virtually all the major riboswitch classes in the ligand-bound state and, for several riboswitches, in the ligand-free state. The resulting spatial topologies have demonstrated the wide diversity of riboswitch folds and revealed structural principles for specific recognition by cognate metabolites. The available three-dimensional information, supplemented by structure-guided biophysical and biochemical experimentation, has led to an improved understanding of how riboswitches fold, what RNA conformations are required for ligand recognition, and how ligand binding can be transduced into gene expression modulation. These studies have greatly facilitated the dissection of molecular mechanisms underlying riboswitch action and should in turn guide the anticipated development of tools for manipulating gene regulatory circuits.
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Biophysical Dynamics in Disorderly Environments
Vol. 41 (2012), pp. 371–402More LessThree areas where time-independent disorder plays a key role in biological dynamics far from equilibrium are reviewed. We first discuss the anomalous localization dynamics that arises when a single species spreads in space and time via diffusion and fluid advection in the presence of frozen heterogeneities in the growth rate. Next we treat the unzipping of double-stranded DNA as a function of force and temperature, a challenge that must be surmounted every time a cell divides. Heterogeneity in the DNA sequence dominates the physics of single-molecule force-extension curves for a broad range of forces upon approaching a sharp unzipping transition. The dynamics of the unzipping fork exhibits anomalous drift and diffusion in a similar range above this transition, with energy barriers that scale as the square root of the genome size. Finally, we describe how activated peptidoglycan strand extension sites, called dislocations in materials science, can mediate the growth of bacterial cell walls. Enzymatically driven circumferential motions of a few dozen of these defects are sufficient to describe the exponential elongation rates observed in experiments on Escherichia coli in a nutrient-rich environment. However, long-range elastic forces transmitted by the peptidoglycan meshwork cause the moving dislocations to interact not only with each other, but also with a disorderly array of frozen, inactivated strand ends.
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Radical Use of Rossmann and TIM Barrel Architectures for Controlling Coenzyme B12 Chemistry
Vol. 41 (2012), pp. 403–427More LessThe ability of enzymes to harness free-radical chemistry allows for some of the most amazing transformations in nature, including reduction of ribonucleotides and carbon skeleton rearrangements. Enzyme cofactors involved in this chemistry can be large and complex, such as adenosylcobalamin (coenzyme B12), simpler, such as S-adenosylmethionine and an iron-sulfur cluster (i.e., poor man's B12), or very small, such as one nonheme iron atom coordinated by protein ligands. Although the chemistry catalyzed by these enzyme-bound cofactors is unparalleled, it does come at a price. The enzyme must be able to control these radical reactions, preventing unwanted chemistry and protecting the enzyme active site from damage. Here, we consider a set of radical folds: the (β/α)8 or TIM barrel, combined with a Rossmann domain for coenzyme B12-dependent chemistry. Using specific enzyme examples, we consider how nature employs the common TIM barrel fold and its Rossmann domain partner for radical-based chemistry.
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Biomolecular Simulation: A Computational Microscope for Molecular Biology
Vol. 41 (2012), pp. 429–452More LessMolecular dynamics simulations capture the behavior of biological macromolecules in full atomic detail, but their computational demands, combined with the challenge of appropriately modeling the relevant physics, have historically restricted their length and accuracy. Dramatic recent improvements in achievable simulation speed and the underlying physical models have enabled atomic-level simulations on timescales as long as milliseconds that capture key biochemical processes such as protein folding, drug binding, membrane transport, and the conformational changes critical to protein function. Such simulation may serve as a computational microscope, revealing biomolecular mechanisms at spatial and temporal scales that are difficult to observe experimentally. We describe the rapidly evolving state of the art for atomic-level biomolecular simulation, illustrate the types of biological discoveries that can now be made through simulation, and discuss challenges motivating continued innovation in this field.
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Previous Volumes
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Volume 53 (2024)
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Volume 52 (2023)
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Volume 51 (2022)
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Volume 50 (2021)
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Volume 49 (2020)
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Volume 48 (2019)
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Volume 47 (2018)
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Volume 46 (2017)
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Volume 45 (2016)
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Volume 44 (2015)
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Volume 43 (2014)
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Volume 42 (2013)
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Volume 41 (2012)
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Volume 40 (2011)
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Volume 39 (2010)
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Volume 38 (2009)
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Volume 37 (2008)
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Volume 36 (2007)
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Volume 35 (2006)
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Volume 34 (2005)
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Volume 33 (2004)
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Volume 32 (2003)
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Volume 31 (2002)
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Volume 30 (2001)
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Volume 29 (2000)
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Volume 28 (1999)
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Volume 27 (1998)
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Volume 26 (1997)
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Volume 25 (1996)
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Volume 24 (1995)
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Volume 23 (1994)
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Volume 22 (1993)
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Volume 21 (1992)
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Volume 20 (1991)
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Volume 19 (1990)
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Volume 18 (1989)
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Volume 17 (1988)
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Volume 16 (1987)
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Volume 15 (1986)
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Volume 14 (1985)
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Volume 13 (1984)
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Volume 12 (1983)
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Volume 11 (1982)
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Volume 10 (1981)
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Volume 9 (1980)
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Volume 8 (1979)
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Volume 7 (1978)
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Volume 6 (1977)
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Volume 5 (1976)
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Volume 4 (1975)
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Volume 3 (1974)
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Volume 2 (1973)
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Volume 1 (1972)
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Volume 0 (1932)