Amino lysozyme

Weber, P. L. Buck, D. R. Capillary Electrophoresis A Past and Simple Method for the Determination of the Amino Acid Composition of Proteins, /. Chem. Educ. 1994, 71, 609-612. This experiment describes a method for determining the amino acid composition of cyctochrome c and lysozyme. The proteins are hydrolyzed in acid, and an internal standard of a-aminoadipic acid is added. Derivatization with naphthalene-2,3-dicarboxaldehyde gives derivatives that absorb at 420 nm. Separation is by MEKC using a buffer solution of 50 mM SDS in 20 mM sodium borate. 

Lysozyme from bacteriophage T4 is a 164 amino acid polypeptide chain that folds into two domains (Figure 17.3) There are no disulfide bridges the two cysteine residues in the amino acid sequence, Cys 54 and Cys 97, are far apart in the folded structure. The stability of both the wild-type and mutant proteins is expressed as the melting temperature, Tm, which is the temperature at which 50% of the enzyme is inactivated during reversible beat denat-uration. For the wild-type T4 lysozyme the Tm is 41.9 °C. 

Just as individual amino acids have isoelectric points, proteins have an overall p/ because of the acidic or basic amino acids they may contain. The enzyme lysozyme, for instance, has a preponderance of basic amino acids and thus has a high isoelectric point (p/= 11.0). Pepsin, however, has a preponderance of acidic amino acids and a low- isoelectric point pi 1.0). Not surprisingly, the solubilities and properties of proteins with different pi s are strongly affected by the pH of the medium. Solubility- is usually lowest at the isoelectric point, where the protein has no net charge, and is higher both above and below the pi, where the protein is charged. 

FIGURE 1119 The lysozyme molecule is a typical enzyme molecule. Lysozyme is present in a number of places in the body, including tears and the mucus in the nose. One of its functions is to attack the cell walls of bacteria and destroy them. This "ribbon" representation shows only the general arrangement of the atoms to emphasize the overall shape of the molecule the ribbon actually consists of amino acids linked together (Section 19.13). 

In most cases the microspheres were insoluble. The polysaccharides might be partially cross-linked via amido groups formed by the carboxyl groups of the polyanion and the restored free amino group of chitosan. The susceptibility to enzymatic hydrolysis by lysozyme was poor, mainly because lysozyme, a strongly cationic protein, can be inactivated by anionic polysaccharides [207]. 

Crystal-structure analysis of Taka amylase A gave similar results, in that it showed that it had an extended cleft which could accommodate six, or possibly seven, a-( 1 — 4)-linked glucose units and two oppositely placed acidic amino acids (Asp-206 and Glu-230) which could interact with the bound substrate similarly to Asp-52 and Glu-35 in lysozyme. 

No sequence homologies can be detected. This is, perhaps, not surprising. The X-ray structure analysis of lysozyme by Phillips has shown that the polypeptide chain is folded in a way which puts none of the amino acids in sequential vicinity of the catalytic Asp-52 and Glu-37 that are near to the bound substrate. Comparable folding patterns can probably be realized with widely differing arrangements of amino acids, and thus the apparent lack of homologies. 

Figure 9 Three-dimensional cationic CITP of (A) blank (B) lysozyme (LYSO), creatinine (CREAT), conalbumin (CAL), y-amino-n-butyric acid (GABA), and ovalbumin (OVA) (C) OVA spiked with CREAT and GABA. Capillary 90 cm (length to the detector, 70 cm) x 75 p i.d. leader 10 mM potassium acetate and acetic acid with 0.3% HPMC, pH 4.75 terminator 10 mM acetic acid sample 10 to 30 mg/ml proteins dissolved in leader without HPMC voltage 20 kV. (From Gebauer, P. and Thormann, W.,. Chromatogr., 558, 423, 1991. With permission.)...

To this list of protein misfolding diseases can be added rare familial amyloidoses in which the mutated proteins have the classic amyloid fibril congophilic birefringence and cross-(3-sheet structure (Table 3). Many of these deposits have an impact on the central nervous system (TTR, cystatin, lysozyme) as well as on other organ systems. A newly described disease, familial British dementia, is associated with the deposition of Abri, a 34 amino acid, 4 kDa peptide cleaved from a 277 amino acid precursor sequence, the last 10 amino acids of which are not normally translated [52]. Familial encephalopathy with neuroserpin inclusion bodies (FENIB) is... 

Enzymes modified with N -carbonyldiimidazole (CDI) include horseradish peroxidase 761 /1-lactamase after nitration and reduction,[771 lysozyme, and urease.[781 Ref. [77] describes how the tyrosine side chain of a protein was nitrated, reduced with dithionite to an amino group, and then treated with CDI or A/-(2,2,2-trifluoro-ethoxycarbonyl)imidazole to give the benzoxazolinonyl alanine moiety ... 

Squaraine dyes 10b, 39a, 39b, 41a, 41c, 41d, and 41e were used to measure different proteins such as BSA, HSA, ovalbumin, avidin from hen egg white, lysozyme, and trypsin (Fig. 12) [58]. It is difficult to predict correlations between the dyes structures and the affinity or sensitivity of the dyes for different proteins. All squaraine probes exhibit considerable fluorescence increases in the presence of BSA. Dicyanomethylene-squaraine 41c is the brightest fluorescent probe and demonstrates the most pronounced intensity increase (up to 190 times) in presence of BSA. At the same time, the fluorescent response of the dyes 10b, 39a, 39b, 41a, 41c, 41d, and 41e in presence of other albumins (HSA and ovalbumin) is, in general, significantly lower (intensity increases up to 24 times). Dicyanomethylene-squaraine 41a and amino-squaraines 39a and 39b are the most sensitive probes for ovalbumin. Dyes 41d, 10b, and 41e containing an A-carboxyalky I -group demonstrate sufficient enhancement (up to 16 times) in the presence of avidin. Nevertheless, the presence of hydrolases like lysozyme or trypsin has only minor effects on the fluorescence intensity of squaraine dyes. 

Strange and Dark demonstrated the presence of a hexosamine containing peptide in the spore coats of B. megaterium and B. subtilis. The breakdown of an insoluble peptide complex might well be one of the first steps of the germination process. It was believed that the release of the hexosamine-amino acid complex was the result of the action of lysozyme present in the spores. 

The hardness of only one type of protein crystal has been found in the literature. It is for lysozyme. This is an enzyme found in egg whites and tears. It destroys bacterial membranes. It is relatively small for a protein molecule, consisting of a chain of 129 amino acids folded into a globule with the volume = 30,000 A3. Its crystals are aggregates of these globular molecules held together by London forces (Stryer, 1988). 

Transition-metal phosphorus trichalcogenides such as MnPS3 are able to intercalate amino acids and peptides by ion exchange. In this way, increases in the basal spacing of 0.7 and 3-4 nm are observed for the intercalation of poly-L-lysine and lysozyme, respectively [224]. Interestingly, the enzymatic activity of the immobilized protein has been detected, suggesting that the enzyme is protected against denaturation. 

Chitosan has found many biomedical applications, including tissue engineering approaches. Enzymes such as chitosanase and lysozyme can degrade chitosan. However, chitosan is easily soluble in the presence of acid, and generally insoluble in neutral conditions as well as in most organic solvents due to the existence of amino groups and the high crystallinity. Therefore, many derivatives have been reported to enhance the solubility and processability of this polymer. 

Habeeb, A.F.S.A., and Atassi, M.Z. (1970) Enzymatic and immunochemical properties of lysozyme. Evaluation of several amino group reversible blocking reagents. Biochemistry 9, 4959—4944. 

Starch (1), Cellulose (2), Cellulose Methyl Ether (3), Oxy-cellulose (4), PVA (5), Partially Hydrolyzed PVAc (6), Silk (.2), Wool (7), Hide-Powder (8), Natural Rubber Latex ( ), Synthesized Poly-(ot-Amino Acids) (10), Nylon-6 (11), Nylon-3 (12), Ot-Amylase (13), Lysozyme (14), RNA (15), Polyacrylonitrile (16), Polyvinyl-sulfonate (17) ... 

An example is provided by the renal delivery of naproxen coupled to lysozyme via an L-lactic acid spacer [261b]. Here, the terminal amino group... 

Lysozym (Abb. 4). Lysoz3un aus HiilinereiweiB cnthalt 129 Amino-saurereste in einer Pol q)eptidkette, welche durch vier Cystinbrucken... 

Some unnatural amino acids have been designed with this metal-chelating property in mind. For instance, bipyridylalanine (BpyAla, 27) has the bipyridyl group that chelates most transition metal ions and has been successfully incorporated into proteins in E. coli BpyAla was shown to reversibly bind copper ions when incorporated into T4 lysozyme, but a tyrosine in the same location was unable to bind copper, indicating that BpyAla is useful to coordinate copper ions to a protein of interest. 

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