Crystallized enzyme proteins

This protein may be further purified, if desired, and eventually may be crystallized, by redissolving the paste in saturated sodium chloride solution by adjusting the pH to 4.5-6.0,and reprecipitating the enzyme protein by the gradual addition of acid in the cold, until a pH of approximately 2.0 is obtained or, the purification may be accomplished by dissolving the protein in acid at a pH of 2, and then precipitating the enzyme, by increasing the concentration of salt. 

Structural information about the oxygenases provided limited insight into the mechanism (Schmidt et al. 2006). The crystallized enzyme from Synechocystis sp. PCC6803 is membrane associated and the interaction with the membrane is believed to be mediated by a nonpolar patch on the surface of the enzyme. This hydrophobic patch is thought to provide the necessary access of the protein to the membrane-bound carotenoids. Following withdrawal from the membrane, the substrate moves through the hydrophobic tunnel toward the metal center. The substrate orients the... 

James Sumner received the Nobel Prize for Chemistry in 1946 for the crystallization of proteins. Richard Willstatter, the 1915 Chemistry prizewinner, had proposed that proteins were not enzymes, and that the protein in urease was simply a scaffold for the veritable catalyst. Since urease is inactive without Ni, he was not so far wrong ... 

The title retains the trivial name for enzymes with the systematic name of urea amidohydrolase and the Enzyme Commission code number of EC 3.5.1.5. Ureases are hydrolases acting on C-N bonds (nonpeptide) in linear amides and thus belong to a group that includes glutaminase, form-amidase, and formyltetrahydrofolate deformylase. The title is plural to emphasize that urease activity may be exhibited by several protein species. Urease, singular, has come to mean by common usage, that particular enzymic protein first crystallized by Sumner from jack bean... 

First and foremost, cross-linked enzyme crystals are crystals. Within the crystal lattice the concentration of protein approaches the theoretical limit. This is important to the process development chemist, who would much rather use a small quantity of a very active catalyst in a reactor than fill it with an immobilized enzyme. Typically an immobilized enzyme contains only 1-10% by weight enzyme, with the remaining carrier material simply occupying valuable reactor space. The crystallinity is absolutely required to achieve the stability exhibited by CLCs [8], Cross-linked soluble thermolysin and cross-linked precipitate of thermolysin are no more stable than the soluble enzyme. Crystals of proteins... 

Hajdu, J., Acharya, K.R., Stuart, D.I., Barford, D., and Johnson, L.N. (1990). Catalysis in enzyme crystals. In Proteins Form and Function. R.A.Bradshaw and M.Purton, eds. (Cambridge Elsevier Trends Journals), pp. 83 93. Hodgson, J. (1990). Protein design mles, empiricism, and nature. Bio/Technology 8,1245 1247. 

Crystal structures of many NHEJ proteins have tremendously supported the discussions above. Also, electron microscopy revealed the overall shape of the large and complex DNA-PKcs structure (23). For most other domains, structural inferences can be drawn by mapping NHEJ proteins onto crystallized enzymes of the same class. However, no published structures show two DSB ends engaged together, which I argue defines the unique functions of NHEJ proteins. Similarly, poor insight exists into the intercoimected assembly of the full NHEJ repair complex. Furthermore, crystal structures are static, whereas NHEJ is dynamic with multiple different reactions. 

Czok, R., and Buecher, T. 1960. Crystallized enzymes from the myogen of rabbit skeletal muscle. Adv. Protein Chem. 15 315-415. 

Mande SC, Mainfiroid V, Kalk KH, Goraj K, Martial JA, Hoi WG. Crystal structure of recombinant human triosephosphate isomerase at 2.8 A resolution. Triosephosphate isomerase-related human genetic disorders and comparison with the trypanosoma enzyme. Protein Sci 1994 3 810-21. 

In contrast to the poor reaction rates reported elsewhere [109], crude powder of lipase from Pseudomonas cepacia (PSL) acylates secondary phenylethyl alcohol at reasonable reaction rates [120] (Table 4). Based on active enzyme protein, the highest reaction rates were observed for carrier-fixed enzymes. This is also true for lipase from Candida antarctica (CAL-B). PSL crystals catalyzed with comparable activity when they had been pretreated with surfactants [109]. 

Immobihzation entails the introduction of inert carrier materials. With the exception of enzyme crystals or enzymes within enzyme membrane reactors, the inert carrier material is usually present in excess of the active enzyme protein. The carrier for crystals is the enzyme protein itself, whose specific activity strictly determines the weight-related activity of the crystal. This is different in the case of dedicated carrier materials. The range of active enzyme can be quite broad because enzyme loading can be adjusted according to the binding capacity of the carrier material. It is therefore possible to establish a well-balanced relationship between reaction volume and carrier by adjusting the amount of bound enzyme on the carrier. 

Cross-linked crystals of lipase from Candida rugosa (CRL) were applied in the resolution of racemic ketoprofen chloroethyl ester. In batch-wise operation, the half-life of the catalyst was reached after about 18 cycles or, in terms of enzyme consumption, about 5.6 g of enzyme protein were consumed to prepare 1 kg of (S)-ketoprofen. CRL suffers from a low specific activity towards this poorly water-soluble substrate which may explain the high enzyme input [117]. 

Nyebar C The crystal is dip coated with a low-surface-energy plastic (Nyebar C 30% solution in l,3-di(trifluoromethyl)benzene). This coating layer is capable of forming hydrophobic bonds with enzymes, proteins, antibiotics, antigens, etc [28]. 

Because the structures of the active sites of enzymes are extremely responsive to the binding of substrates and substrate analogues, conformational changes may take place which give rise to cracking or disorder of the crystal. It is important, however, that the crystals of protein plus inhibitor be closely isomorphous with the native protein crystals, at least with respect to the cell dimensions so that the molecular transform is sampled at the same reciprocal lattice points. In those instances where large conformational changes occur, de novo structure determination has to be attempted. 

Pale, btownish-green Crystals or shiny, transparent, striated plates. Practically insol in water. Slightly sol in buffered alkali solns. So] ns at pH 7.5-10.5 are relatively si able. Shows unusually high absorption in the region of 35777350 nm (for highly purified uricase AjS = 11-3, AJJj — 2.0, both in 1% Na,CO, Nm/Aija = 5.6). Isoelec, pt, pH 6.3. Copper content of the enzyme (specific activity [20-125) equals 0,56 mg/g of enzyme protein. There appears to be a satisfactory correlation between copper content and activi-iy of the purified enzyme. The enzyme is sensitive to cyanide ion the ptesence of I0-4M KCN inhibits its activity. 

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