Extracellular enzymic analysis

Digestion of PGA by the PelL enzyme yielded a mixture of unsaturated ohgogalacturonides, giving evidence that PelL is an endo-deaving lyase (17). An exo-enz3mie, such as the EC 16 PelX, would generate a single product (15). The PelL protein differs from the major E. chrysanthemi pectate lyases in its ability to cleave both PGA and methylated pectin (17). The PelL activity has a basic optimum pH and an absolute requirement for Ca + ions. Analysis of culture supernatants demonstrated that PelL is an extracellular enzyme, such as the other secondary pectate lyases (17). 

Extracellular functions of calcium are better understood than intracellular functions, partly because of the difficulties associated with the analysis of calcium inside the cell. Extracellular roles are those of (a) trigger, (b) an activator and stabilizer of extracellular enzymes, and (c) in bone and teeth (Section 62.1.3.9). Intracellular activities are mainly trigger effects, and are intimately associated with problems of transport and binding. 

E. Zilkha, T.P. Obrenovitch, A. Koshy, H. Kusakabe, and H.P. Bennetto, Extracellular glutamate online monitoring using microdialysis coupled to enzyme-amperometric analysis. J. Neurosci. Methods. 60,1-9 (1995). 

Neurotransmitters, peptides, and other endogenous substances in the extracellular space can be sampled using in vivo microdialysis. The dialysis membrane typically excludes the transport of larger molecules and enzymes that could otherwise interfere with analysis of the substances of interest. In the case of neurotransmitters, the levels in dialysates are the net result of the interaction between processes affecting release into and removal from the extracellular space. Consequently, in vivo microdialysis can only sample a neurotransmitter that has not yet been removed by clearing mechanisms and this method does not provide information regarding intracellular levels of substances. 

The extracellular polysaccharides of Rhizobium meliloti 201 have been examined by using enzymic degradation and chemical procedures.314 A mixture of polysaccharides produced by the bacterium, when incubated with a bacterial enzyme that hydrolyzed one of these, gave oligosaccharides that could be separated by DEAE-cellulose chromatography. The major fraction was a pentasaccharide, for which methylation analysis and Smith... 

The sequence analysis of the /3-lactamase protein of B. licheniformis 749/C was performed on both the extracellular and the cell-bound enzyme (of. serial Nos. 5 and 6, Table I). The slight differences between the two forms were confined to the N-terminus and attributed to the difference in the release mechanism. Peptides corresponding to about 90% of the molecule have been characterized and their sequences combined to form five larger fragments. 

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