bluuebunny/crossref_metadata_2025_split huggingface dataset:
High-throughput combinatorial synthesis of discrete compounds in multimilligram quantities: non-chemical encoding and directed sorting
Authors: Xiao* Xiao-yi , Nicolaouꝉ K C 
Published: November 2023

Abstract
Over the past few years the pharmaceutical community’s need to generate and screen chemical libraries with large diversity has fuelled the rapid development of combinatorial chemistry (1, 2). Research areas such as high-throughput solution (3, 4) and solid phase (5-7) synthesis, the development of novel solid supports and linkers (Chapters 6 and 7) (8, 9), as well as new powerful encoding strategies (10-14) (Chapter 1) have received exceptional attention due to their central role in the development of this field. The chemical encoding strategies (15, 16), coupled with the split and pool technique (17) (Chapters 1-3) greatly increase the efficiency of combinatorial synthesis and screening. However, these strategies produce each member of the library in very small amounts (nanograms to micrograms), making subsequent multiple screenings and characterization procedures quite difficult. Although traditional parallel synthesis (18, 19) in combination with the remarkable recent progress in laboratory automation (19) produces larger quantities of material, it still lacks the efficiency of the split and pool technique particularly for the preparation of very large libraries. We have recently developed two new combinatorial synthesis strategies employing non-chemical encoding and directed sorting in a batch synthesis fashion (10, 11) that combine both the advantages of split and pool and parallel synthesis. These strategies are capable of producing large combinatorial libraries and delivering milligram quantities of each library member. This chapter discusses the development of these two non-chemical encoding techniques: radiofrequency (Rf) tagging (10) and bar coding (11) in combination with a directed sorting strategy, and their application to the synthesis of combinatorial libraries of a variety of chemical structures.
Combinatorial synthesis on multivalent oligomeric supports
Authors: Kim Ronald M , Chang Jiang 
Published: November 2023

Abstract
This chapter describes a strategy for liquid phase combinatorial chemistry which we term combinatorial synthesis on multivalent oligomeric supports (COSMOS). In this approach, compounds are synthesized on multivalent soluble supports through iterative cycles of solution phase synthesis and size-based product purification. Final products may be cleaved from the support, and isolated from the oligomer by size. A brief comparison of solid phase and liquid phase synthesis will be presented, followed by an introduction to COSMOS methodology. The role of the support in COSMOS, and methods for rapid product purification will be highlighted. Detailed procedures for the preparation of soluble supports, and an illustrative synthesis using the COSMOS approach will be described. Emphasis will be placed on protocols for isolating support-bound and cleaved products. Parallel sample processing and automation of the COSMOS methodology will be discussed.
Differential display analysis of memory-associated genes
Authors: Huang A-Min , Lee Eminy H Y 
Published: November 2023

Abstract
Memory formation is a complicated process, especially in mammals. In the form of long-term memory formation, it is well known that it requiresde novaRNA and protein synthesis because protein and mRNA synthesis inhibitors impair long-term memory formation (1, 2). In the invertebrate system, extensive efforts have been made to identify specific genes associated with memory formation. For example, by using quantitative 2D gel analysis, Castellucciet al.(3) have identified several proteins that are related to the process of long-term sensitiza tion, a form of long-term memory studied in the invertebrateAplysia.Also, screening ofDrosophilamutants has yielded about ten genes that are related to the process of olfactory learning and memory (4). Although these methods are effective, two disadvantages exist by using these approaches for studies in the mammalian system. First, they take a long time to identify and characterize the genes of interest. Secondly, gene expression associated with learning and memory in mammals is much more complicated which may not be well detected by using the traditional methods.
Glycosylphosphatidylinositol-anchored proteins and their phospholipases
Authors: Low Martin G 
Published: November 2023

Abstract
A large number of eukaryotic cell-surface proteins are covalently linked to glyco-sylphosphatidylinositol (GPI). A GPI molecule consists of a short glycan chain containing four sugar residues glycosidically linked to the 6-OH of a phosphatidylinositol molecule (see Fig. 1). The other, non-reducing end of the glycan chain bears a phosphodiester-linked ethanolamine residue. A GPI molecule is attached post-translationally to proteins via an amide linkage between the terminal phospho-ethanolamine and the carboxyl group of the C-terminal amino acid. The lipid moiety of the GPI is inserted into the lipid bilayer and is responsible for anchoring the protein to the cell membrane. Proteins can utilize other types of lipid anchor, involving prenyl, palmitoyl, or myristoyl groups but, in eukaryotic cells, GPI is the only lipid that can anchor proteins to the cell surface. The GPI anchor is susceptible to cleavage by several phospholipases that are distinct from those involved in the hydrolysis of intracellular inositol phospholipids. Cleavage of the anchor by these phospholipases releases GPI-anchored proteins from the membrane. The relatively complex structure of the GPI molecule and evolutionary conservation of its major features suggest that the GPI anchor confers additional properties on the proteins besides attachment to the cell surface. GPI molecules and GPI-anchored proteins have been proposed to play a role in cellular signalling as well as targeting proteins to particular regions of the cell surface. However, the mechanism and physiological significance of these additional functions is uncertain.
Assessment of DNA damage cell cycle checkpoints in G1 and G2 phases of mammalian cells
Authors: O’connor Patrick M , Jackman Joany 
Published: November 2023

Abstract
Cell cycle progression is normally tightly regulated such that one cell cycle event does not occur unless the previous cell cycle event has been completed. For example, mammalian cells do not normally enter mitosis while DNA replication is on-going, and cells in mitosis do not normally exit mitosis unless the spindle apparatus is correctly aligned with all the chromosomes. This dependency is controlled by a series of negative-feedback control systems that are called checkpoints (1-4). These checkpoints are being studied in a variety of organisms from yeast to human, and a growing number of studies implicate defective checkpoint control systems in the emergence and evolution of human cancer. One example is the p53 tumour suppressor, the most commonly mutated gene in human cancer (1, 2). p53 is essential for the arrest of cell cycle progression in the 01 phase following DNA damage. Loss of p53 function prevents this arrest mechanism from being activated and also sub dues or blocks the induction of apoptosis in DNA damaged cells. In addition, p53 has influences on other cell cycle checkpoints (02 and M phase) which act together to ensure the fidelity of DNA replication and chromosome segregation. Such actions help to explain why there is a markedly increased likelihood of cancer in mice deficient for p53 function (5).
Human leukaemia cells as a model differentiation system
Authors: Moore Dorothy C , Studzinski George P 
Published: November 2023

Abstract
Cell differentiation usually proceeds at the same time as cessation of cell proliferation, andin vitrosystems for cell differentiation provide excellent models to study orderly down-regulation of growth-related cellular activities. Many such models are available, and these are illustrated in this volume with reference to systems frequently used to study differentiation of human malig-nant cells. Differentiation of colon cancer cells is described in Chapter 12, of human prostate cancer in Chapter 13, and of human leukaemia in this chapter.
Active microelectronic arrays for DNA hybridization analysis
Authors: Heller Michael j. , Tu Eugene , Holmsen Anita 
Published: November 2023

Abstract
Miniaturized DNA arrays, DNA chips, and lab on a chip devices represent a new area of high technology that will revolutionize the way many important molecular biological-type experiments, assays, and tests are performed in both research and clinical diagnostic laboratories. The development of these new technologies has been brought about by a synergistic combination of diverse disciplines which include: microfabrication, organic chemistry, molecular biology, and genetics. In particular, many of these devices and technologies take advantage of sophisticated microfabrication processes developed by the semiconductor industry over the last 40 years.
Assaying binding and covalent modifications of cytoskeletal proteins
Authors: Ampe Christophe , Vandekerckhove Joël 
Published: November 2023

Abstract
Cytoskeletal dynamics are essential for cell survival. The microfilament system is one of the major players in cell motility processes. It exerts force for movement either by the action of molecular motors on relatively stable filaments or by dynamic turnover of actin filaments. In the latter case processes such as filament polymerization, depolymerization. and bundling are regulated in a temporal and spatial manner, governed by actin-binding proteins. The complexity of the actin system arises from the multitude of actin-binding proteins, often with partly overlapping activities, displaying multiple functions or effects on actin polymerization, depending on the intracellular environment. Therefore, detailed biochemical studies of individual actin-binding proteins and their interaction with actin or with regulatory ligands are necessary to understand cellular motility phenomena. The field is now slowly progressing towards in vitro studies with more than one actin-binding protein, because it is dear that in cells these proteins either compete with each other or have additive or co-operative effects on aetin reorganization.
Ras-related GTPases and the cytoskeleton
Authors: Ridley Anne J , Flinn Helen M 
Published: November 2023

Abstract
The Ras supcrfamily or GTPascs all hind GTP, and can hydrolyse GTP to yield Ci DP. They act as molecular witches in cells and are active when bound to GTP, assuming a conformation which allows them to interact with their downstream target proteins. In the GDP-bound conformation they no longer hind to most target proteins (I, 2). Under physiological conditions, exchange of guanine nucleotides on the GTPases is very slow and is catalysed by exchange factors, while GTP hydrolysis is catalysed by GTPase activating proteins (GAPs) (Figure 1).
Overview of apoptosis
Authors: Studzinski George P 
Published: November 2023

Abstract
When cells receive mixed signals for growth they usually die. For instance, when the developmental programme requires cell division but external growth signals are lacking, or when a growth-related gene such as c-myc is highly expressed but the cellular environment has insufficient nutrient content, or a toxic xenobiotic is present, the cell dies by a process termed apoptosis. Although there are differences in the phenomena observed during the apoptotic sequence of events, depending on the cell type, and agent or circumstance which initiates the cell’s demise, there are morphological and biochemical similarities which suggest that these are variants of the same biological process, designed to control the size of cell populations.
Sphingolipids as messengers of cell death
Authors: Jenkins Gary M , Hannun Yusuf A 
Published: November 2023

Abstract
This chapter discusses the most commonly used methods for the study of sphingolipids in their role as messengers of cell death. A basic review of sphingolipids and the role they play in apoptosis is given. The rest of the chapter outlines the methods used for the study of sphingolipids and the enzymes relevant to the sphingomyelin (SM) cycle. Each method is described with full experimental details, and emphasis is given to one or two protocols used for the study of each class of sphingolipids and relevant enzymes.
Multiple alignments for structural, functional, or phylogenetic analyses of homologous sequences
Authors: Duret L , Abdeddaim S 
Published: November 2023

Abstract
Understanding the structure, function and evolution of genes is one of the main goals of genome sequencing projects. Classically, gene function has been investigated experimentally through the analysis of mutant phenotypes. More recently, comparative analysis of homologous sequences has proved to be a very efficient approach to study gene function (this approach has been coined ‘comparative genomics’ or ‘phylogenomics’). Indeed, the evolution of living organisms maybe considered as an ongoing large-scale mutagenesis experiment. For more than three billion years, genomes have continuously undergone mutations (substitutions, insertion, deletions, recombination, and so on). Deleterious mutations are generally rapidly eliminated by natural selection, while mutations that have no phenotypic effect (neutral mutations) may, by random genetic drift, eventually become fixed in the population. Globally, advantageous mutations are very rare, and hence residues that are poorly conserved during evolution generally correspond to regions that are weakly constrained by selection (l). Thus, studying mutation patterns through the analysis of homologous sequences is useful not only to study evolutionary relationships between sequences, but also to identify structural or functional constraints on sequences (DNA, RNA, or protein).
Introduction to basic principles
Authors: Freshney R Ian 
Published: November 2023

Abstract
Cell culture has become an indispensable technology in many branches of the life sciences. It provides the basis for studying the regulation of cell proliferation, differentiation and product formation in carefully controlled conditions, with processes and analytical tools which are scalable from the level of the single cell to in excess of 10 kg wet weight of cells. Cell culture has also provided the means to define almost the entire human genome, and to dissect the pathways of intracellular and intercellular signalling which ultimately regulate gene expression. From its ancestry in developmental biology and pathology, this discipline has now emerged as a tool for molecular geneticists, immunologists, surgeons, bioengineers, and manufacturers of phannaceuticals, while still remaining a fundamental tool to the cell biologist, whose input is vital for the continuing development of the technology.
Cell polarity in yeast
Authors: BÄHler JÜrg , Peter Matthias 
Published: November 2023

Abstract
Eukaryotic cells respond to intracellular and extracellular cues to direct cell growth and division. Selecting sites of polarized growth and division is crucial for the development of both unicellular and multicellular organisms. The two yeasts Saccharomyces cerevisiae (budding yeast) and Schizosaccharomyces pombe (fission yeast) have rigid cell walls. Cellular polarization targets secretion of cell wall and other materials to restricted areas in the cell. As a result, cell polarity in these yeasts directly underlies cellular morphogenesis. Both budding and fission yeast establish cell polarity at several stages for growth, cytokinesis, mating, and sporulation. Thus, they are useful model organisms for studying various aspects of cell polarity. In recent years, many regulatory and cytoskeletal components important for directing and establishing polarity have been identified, and molecular mechanisms underlying polarity development in yeast have been elucidated. Key signalling pathways that regulate polarization during the cell cycle and mating response have been described. Since many of the components important for polarized cell growth are conserved in other organisms, the basic mechanisms mediating cell polarity are likely to be universal among eukaryotes.
Cell polarity in algae and vascular plants
Authors: Fowler John E 
Published: November 2023

Abstract
Due to the presence of the plant cell wall, the morphogenesis and physiology of plant cells and animal cells are under different constraints. Plant cells are immobile, and fixed in relation to their neighbours; morphogenetic changes in the plant cannot occur by the cellular migration mechanisms common during animal development. Furthermore, modificati6n of plant cell shape must occur through modification of the surrounding cell wall. Thus, mechanisms that allow co-ordination and expression of morphogenetic changes in plants are coupled with mechanisms that regulate growth, both by expansion and division of cells.
Genetic analysis of intrinsically asymmetrical cell division
Authors: Piano Fabio , Kemphues Kenneth 
Published: November 2023

Abstract
Intrinsically asymmetrical cell division is a major mechanism for generating cell diversity. In this process, referred to simply as asymmetrical division in this chapter, cytoplasmic and cortical asymmetries within a mother cell combine with an oriented cell division to produce daughter cells with different cytoplasmic components and consequently different cell fates (1). As illustrated in Fig. 1, there are three processes that contribute to asymmetrical cell divisions: (1) establishment of polarity; (2) localization of determinants; (3) oriented cell division.
Polarized exocytosis: targeting vesicles to specific dotnains on the plastna membrane
Authors: Brennwald Patrick , Adamo Joan 
Published: November 2023

Abstract
The ability to deliver newly synthesized proteins and lipids to discrete sites on the cell surface allows cells to grow asymmetrically (i.e. tall and thin versus round), to release morphogens on a discrete side of a cell layer during early embryonic development, and to carry out a variety of other processes. The involvement of delivery events is particularly apparent during asymmetrical growth. This is because the ability of the surface to expand requires the addition of new phospholipids and integral membrane proteins, which are the building blocks that allow cell-surface growth. Lipids and integral membrane proteins are extremely hydrophobic in nature and thus they cannot reach the plasma membrane by simple diffusion through the cytoplasm. Instead, they are brought to the cell surface in the form of membranous transport vesicles. These transport or secretory vesicles contain materials first synthesized in the endoplasmic reticulum, and then delivered to the Golgi apparatus, within which the secretory vesicles themselves are derived. These post-Golgi secretory vesicles represent the last stage of the exocytic pathway and their fusion with the cell surface is often referred to is exocytosis. In this chapter, we will discuss the mechanisms and machinery that regulate the delivery and fusion of secretory vesicles with specific sites on the plasma membrane, the process of polarized exocytosis.
Determination of active site concentration
Authors: Martin Mark T 
Published: November 2023

Abstract
Enzyme quantitation methods vary widely in complexity and reliability (Table 1). Methods that assume an enzyme is homogeneous and fully active (gravimetric or total protein assays) are far less reliable than methods that measure the concentration of functional enzyme (active site titrations1 and substrate turnover rate assays). Of those methods that measure functional enzyme, active site titrations tend to be more reliable than activity assay methods. In contrast to activity assay methods, titrations do not require primary standards of 100% pure and active enzyme and are less likely to be affected by small variations in assay conditions (pH, ionic strength, temperature, etc.).
High throughput screening – considerations for enzyme assays
Authors: Hayes David , Mellor Geoff 
Published: November 2023

Abstract
The rapidly increasing number of potential therapeutic targets is leading the pharmaceutical industry to radically alter its drug discovery strategy (for reviews, see 1–6). For these targets, there is often little or no structural information available for the rational design of putative drug candidates. Additionally, there is a desire to obtain ‘lead’ molecules of novel structure, to avoid patent infringement of known inhibitors. One approach to address these issues is to assay large numbers of compounds against the target, either in a random or sometimes targeted fashion, a process termed high throughput screening (HTS). The precise nature of these targets varies widely; for example they can be receptor–ligand, protein–protein, protein–carbohydrate, DNA and RNA interactions, ion-channels or enzymes. For this chapter we will consider only enzyme targets and will discuss the requirements for successful assay development and validation and subsequent screening. Details of two example assays are given, along with a snapshot of automation required to facilitate the process.
Statistical analysis of enzyme kinetic data
Authors: Cornish-Bowden Athel 
Published: November 2023

Abstract
All serious kinetic investigations of enzymes include some data analysis, and the methods used can be classified into three categories: (i) graphical analysis; (ii) best-fit analysis with general-purpose commercial packages; (iii) best-fit analysis with specialized programs designed with enzyme kinetic experiments in mind. The near-universal availability of small computers—a major change from the context in which the chapter corresponding to this one in the first edition of this book (1) was written—has greatly increased the use of the second of these approaches, but the others continue to be widely used.
Gel electrophoresis of proteins
Authors: Gartin David E 
Published: November 2023

Abstract
At one time or another during the course of protein analysis or purification, researchers are likely to make use of gel electrophoresis. All laboratories working with proteins have some capability for carrying out gel electrophoresis and all researchers have at least rudimentary knowledge of the technique. Gel electrophoresis can provide information about the molecular weights and charges of proteins, the subunit structures of proteins, and the purity of a particular protein preparation. It is relatively simple to use and it is highly reproducible. The most common use of gel electrophoresis is the qualitative analysis of complex mixtures of proteins. Microanalytical methods and sensitive, linear image analysis systems make gel electrophoresis popular for quantitative and preparative purposes as well.
Fluorescence-based iber-optic biosensors
Authors: Walt David R , Schauer Caroline L , Stitzel Shannon E , Fleming Michael S , Epstein Jason R 
Published: November 2023

Abstract
This chapter describes optical biosensors that combine analyte-sensitive indicators with optical fibers for detecting chemical and biological species. Fiber-optic biosensors are more versatilebecause they are flexible in terms of their platform design and signal transduction. Optical transduction mechanisms include fluorescence, absorbance, and reflectance. Using these mechanisms, sensors can be designed to exploit optically detectable differences in intensity, lifetime, spectral shape, polarization, and wavelength (1). Optical fibers come in a range of lengths and diameters, and can be adapted for either single-or multianalyte detection, and can incorporate a variety of sensing chemistries.