Probes of Biological Processes

Non-canonical Amino Acids as Probes of Biological Processes. 206... 

With these guidelines in mind, specific approaches to probe development can be implemented to transition forward chemical genetic screening hits into validated probes of biological processes, and these strategies are highlighted in Subheading 3. 

The power of the pooled GST fusion protein approach will increase as new biochemical reagents and assays become available. The development of chemical probes for biological processes, termed chemical biology, is a rapidly advancing field. For example, the chemical synthesis of an active site directed probe for identification of members of the serine hydrolase enzyme family has recently been described (Liu et al., 1999). The activity of the probe is based on the potent and irreversible inhibition of serine hydrolases by fluorophosphate (FP) derivatives such as diisopropyl fluorophosphate. The probe consists of a biotinylated long-chain fluorophosphonate, called FP-biotin (Liu et al., 1999). The FP-biotin was tested on crude tissue extracts from various organs of the rat. These experiments showed that the reagent can react with numerous serine hydrolases in crude extracts and can detect enzymes at subnanomolar... 

Barker, A.L., Slevin, C.J., Unwin, P.R. and Zhang, J. (2001) Scanning electrochemical microscopy as a local probe of chemical processes at liquid interfaces. Chapter 12 in A.G. Volkov (Ed.) Liquid Interfaces in Chemical, Biological and Pharmaceutical Applications. Dekker, New York. See also Chapter 6. 

Sialic acids are involved in a number of biological processes including cell-to-cell, cell-to-microorganism, -toxin, and -antibody binding. Their importance in these processes, especially those with relevance to human disease states, has led to interest in the synthesis of both natural and modified sialic acids. This review examines the most recent methods used for the synthesis and modification of sialic acids and for the preparation of sialyl glycosides as biological probes of sialic acid-recognising proteins. 

The synthesis of enantiomers of natural products is a field of increasing interest because such compounds are used in structural studies and as probes for the elucidation of biological processes, as e.g. exemplified by the synthesis of e f-enterobactin [134]. We have reported on the synthesis of (2iS)-2-P-L-glucopyranosyloxy-4-hydroxy-7-methoxy-2//-l,4-benzoxazin-3(4//)-one (e f-GDIMBOA) starting from L-glucose and DIMBOA [135] following the method described in Fig. (18) on principle. 

The examples in this section have been chosen to provide an in-depth presentation showing how RSSF currently has been applied to the study of biological systems. These applications include the study of isotope effects on enzyme-catalyzed reactions, the investigation of substrate-metal ion interactions in metalloenzymes, the search for and identification of covalent intermediates in enzyme-catalyzed processes, the analysis of the effects of site-directed mutations on enzyme catalytic mechanism, and the exploitation of natural and artificial chromophores as probes of allosteric processes. 

The detailed knowledge of the different steps of biological processes on a molecular level is one of the ambitious goals of molecular biology. The importance of this field was underlined by the award of the Nobel Prize in chemistry in 1988 to J. Deisenhofer, R. Huber, and H. Michel for the elucidation of the primary steps in photosynthesis and the visual process [1511]. This subsection illustrates the importance of time-resolved Raman spectroscopy in combination with pump-and-probe techniques (Sect. 6.4) for the investigation of fast biological processes. 

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