Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.24.27 (thermolysin)
 1,894 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Despite the well-characterized function of the green-algal eyespot apparatus as a combined absorption/reflection screen for the photoreceptor for phototaxis, little is known about the proteins involved in the formation of this complex organelle. We therefore purified the carotenoid-rich lipid globules, which are the most conspicuous component of the eyespot sensu strictu from Spermatozopsis similis Preisig et Melkonian. Electron microscopy and an average carotenoid:chlorophyll ratio of 51, confirmed the high purity of the fraction. The diameter of isolated globules (approx. 112 nm) fell within their in vivo range (90-120 nm). Absorption spectra in aqueous media peaked at 535 nm. The predominant carotenoids were beta/psi-, beta, beta- and delta-carotene. Freeze-fracture studies with cells and whole-mount electron microscopy of isolated globules demonstrated regularly arranged particles at the globule surface. Sodium dodecyl sulfate polyacrylamide gel electrophresis revealed specific enrichment of 10 tightly bound major proteins and several minor proteins with the globules. Proteases were used to analyze their topology and function. Upon treatment with thermolysin, globules were released from a fraction enriched in isolated eyespot apparatuses. Major proteins of these globules, and those treated with thermolysin after isolation, were identical. However, the purified proteins were sensitive to thermolysin, indicating that domains of them are normally hidden in the globule matrix. In contrast, pronase degraded all globule-associated proteins in situ. These globules were not stable and easily fused, whereas thermolysin-treated globules were relatively stable. Lipase did not affect globule stability. These results indicate that the five thermolysin-resistant proteins (apparent Mr values: 56, 52, 32, 29, 27 kDa) are close to the surface and might be crucial for globule stabilization, whereas the thermolysin-accessible proteins are probably involved in globule/globule interactions and/or globule/eyespot-membrane interactions.
 ...
PMID:Subfractionation of eyespot apparatuses from the green alga Spermatozopsis similis: isolation and characterization of eyespot globules. 1152 56

Freeze-drying (lyophilization) removes water from a frozen sample by sublimation and desorption. It can be viewed as a three-step process consisting of freezing, primary drying and secondary drying. While cryoprotectants can protect the protein from denaturation during early stages, lyoprotectants are needed to prevent protein inactivation during drying. The structural changes as a result of freeze-drying have been investigated, especially by FTIR (Fourier-transform IR) spectroscopy. In general, drying results in a decrease of alpha-helix and random structure and an increase in beta-sheet structure. In the case of basic fibroblast growth factor and gamma-interferon, enhanced FTIR showed large conformational changes and aggregation during freeze-drying, which could be prevented by using sucrose as a lyoprotectant. It is now well established that structural changes during freeze-drying are responsible for low activity of freeze-dried powders in nearly anhydrous media. Strategies such as salt activation can give 'activated' enzyme powders, e.g. salt-activated thermolysin-catalysed regioselective acylation of taxol to give a more soluble derivative for therapeutic use. In the presence of moisture, freeze-dried proteins can undergo disulphide interchange and other reactions which lead to inactivation. Such molecular changes during storage have been described for human insulin, tetanus toxoid and interleukin-2. Some successful preventive strategies in these cases have also been mentioned as illustrations. Finally, it is emphasized that freeze-drying is not an innocuous process and needs to be understood and used carefully.
 ...
PMID:Freeze-drying of proteins: some emerging concerns. 1503 37