T4 phage lysozyme

Myoglobin, hemoglobin Thermolysin domain 2 T4 phage lysozyme domain 2 Papain domain 1... 

Tyrosyl-tRNA synthetase dl, d2 Thermolysin dl, d2 T4 phage lysozyme dl, d2 Glucosephosphate isomerase dl, d2 Pyruvate kinase dl, d2 Pyruvate kinase d2, d3 Lactate dehydrogenase dl, d2 Alcohol dehydrogenase dl, d2 Glyceraldehyde-phosphate dehydrogenase dl,d2... 

Matthews, B. W. (1975). Comparison of the predicted and observed secondary structure of T4 phage lysozyme. Biochim Biophys Acta 405,442-51. 

B. W. Matthews, Biochim. Biophys. Acta, 405, 442 (1975). Comparison of Predicted and Observed Secondary Structure of T4 Phage Lysozyme. 

Lysozymes derived from sources other than bird egg-whites were also tested for their susceptibility to inactivation by the epoxypropyl /8-gly-cosides. Among these, human leukemic urine lysozyme was inactivated by (GlcNAc)3-Ep at pH 5.5, whereas the lysozymes from papaya latex and the bacteriophage T4 were not inactivated at all. Similar results were also obtained at pH 4.6, at which the papaya lysozyme acts on chitin most efficiently. These results show that the active sites of human and the hen lysozymes are similar, whereas the active sites of the T4 phage and the papaya lysozymes differ from them. Indeed, the hen and the human lysozymes are similar in their amino acid sequences" and three-dimensional structures, whereas the papaya and the T4 phage lysozymes differ from the hen egg-w hite lysozyme in their amino acid composition, molecular weight, and substrate specificity."" ... 

Singla et al. [19] examined the adsorption kinetics of wild type and two synthetic stability mutants of T4 phage lysozyme at silanized silica surfaces. Substitution of the isoleucine at amino acid position three with cysteine (13 C) and tryptophan (13W) rendered such mutants with a higher and lower thermal stability, respectively. It was found that the I3W mutant, characterized by a lower structural stability, would more readily undergo a stmctural change at the interface. Moreover, such a mutant showed more resistance to elution by DTAB than either the wild type or 13 C mutant, simply by forming a more tightly bound conformation (adsorbed state) with the adsorbent. 

The above properties have enabled us to investigate the biochemical properties of the two forms of the lipoprotein and their quantitative relationship. Figure 8 shows a gel pattern of the E. coli envelope proteins after T4 phage lysozyme treatment. The only change caused by the treatment is the appearance of a new peak at the arrow in Fig. 8. Without lysozyme treatment, no such peak appears, as shown in Figs. 2 and 4. All murami-dases tested so far (hen egg-white lysozyme, goose egg-white lysozyme, and T4 phage lysozyme) cause the same peak to appear. ... 

Fig. 8. Gel electrophoresis of an envelope protein fraction double labeled with pHJarginine and [ CJhistidihe. The envelope fraction was treated with T4 phage lysozyme before electrophoresis.

Fig. 10. Pulse-chase experiment of the envelope fraction. (A) A culture was pulse labeled for 4 min with [ C]arginine. Another culture was pulse labeled for 4 min with PH]arginine and chased with nonradioactive arginine for another 50 min. The envelope fraction was prepared from the mixture of both cultures, digested with T4 phage lysozyme, and subjected to SDS gel electrophoresis on a 7.5% acrylamide gel. (B) The radioisotopes were used in the reverse way...

Fig. 11. Gel electrophoresis of the envelope fractions of an E. coli histidine auxotroph ( . coli CP78) labeled with [ C]arginine in the presence and absence of histidine. (A) Labeled in the presence of histidine for 1 h. (B) Labeled during a 1-h histidine starvation. (C) Labeled for 2 h after 2 h of histidine starvation. The patterns of the envelope fractions with and without the lysozyme treatment were superimposed with the aid of the internal standards. Envelope fractions treated with T4 phage lysozyme ------- without T4 phage treatment . In the case of...

A T4 bacteriophage lysozyme (from phage grown on Eco/ZB ) [12585-29-4] was extracted and freed from... 

Exit of the virus from the cell occurs as a result of cell lysis. The phage codes for a lytic enzyme, the T4 lysozyme, which causes an attack on the peptidoglycan of the host cell. The burst size of the virus (the average number of phage particies per cell) depends upon how rapidly lysis occurs. If lysis occurs early, then a smaller burst size occurs, whereas slower lysis leads to a higher burst size. The wild type phage exhibits the phenomenon of lysis inhibition, and therefore has a large burst size, but rapid lysis mutants, in which lysis occurs early, show smaller burst sizes. 

Polysaccharide chains in the peptidoglycan layer (Fig. 8-29) of the cell walls of bacteria are attacked and cleaved by lysozymes,55 enzymes that occur in tears and other body secretions and in large amounts in egg white. Some bacteria and fungi, and even viruses, contain lysozymes.56 Their function is usually to protect against bacteria, but lysozyme of phage T4 is a component of the baseplate of the virus tail (Box 7-C). 

Since there are strict stereochemical requirements for the relative positions and orientations of the two participating cysteine residues,11 addition of new disulfides to existing proteins by site-directed mutagenesis has not always produced the desired increase in stability. Introduction of disulfide bonds has been attempted for phage T4 lysozyme,4-71 phage A repressor,81 dihydrofolate reductase,91 and subtilisins.10-131 Among them the most extensive study has been performed on T4 lysozyme, and enhancement of protein stability has been successful. 

Nicholson, H., Anderson, D.E., Dao-Pin, S., and Matthews, B.W. (1991) Analysis of the interaction between charged side chains and the a-helix dipole using designed thermostable mutants of phage T4 lysozyme, Biochemistry 30, 9816-9828. 

Table 19.9. Thermodynamic stability8 of phage T4 lysozymes with different amino acids at position 157 [646]...

In contrast, in phage T4 lysozyme a thermolabile mutant was found in which one single amino acid was exchanged Thr- Ile 157, and the thermal melting point dropped from 42°C for the wild-type enzyme to 31 °C for the mutant. Systematic exchange of the amino acid in position 157 by site-directed mutagenesis led to the... 

Fig. 19.19. Schematic description of the hydrogen-bonding geometry formed with different amino acids at position 157 in phage T4 lysozyme. The potential hydrogen-bond lengths 1,2,3 and associated angles a, p, y are ...

It becomes clear from this study that hydrogen bonding is of prime importance, although other effects due to van der Waals interactions and steric repulsion contribute when amino add 157 in phage T4 lysozyme is mutated. The deleterious influence of lie 157 in the thermosensitive mutant can be explained on this basis, as well as the effects of mutations where Thrl57 is substituted by Phe, His, Val (see Thble 19.9). In these cases, all hydrogen bonds in the pocket around the side-chain of amino acid 157 are broken except for one to a water molecule, Thr 1550 y Ow. 

Alber T, Dao-pin S, Wilson K, Wozniak JA, Cook SP, Matthews BW (1987) Contributions of hydrogen bonds of Thr 157 to the thermodynamic stability of phage T4 lysozyme. Nature (Lond) 330 41-46... 

Pjura P, Matsumura M, Baase WA, Matthews BW. Development of an in vivo method to identify mutants of phage T4 lysozyme of enhanced thermostability. Protein Sci. 1993 2 2217-2225. 

Sites within the carboxy-terminal domain core of phage T4 lysozyme were substituted singly and as a group with methionine to produce a simplified core sequence. The properties of such mutant lysozymes are briefly described. In addition we describe a method to isolate mutant protein from inclusion bodies and a sensitive enzymatic assay to detect small differences in mutant protein activities. 

The fact that at least seven core residues can be replaced as a group with methionine in phage T4 lysozyme without introducing molten globule-like characteristics shows that strict side-chain complementarity is not required to maintain native-like protein properties. 

Mann, G., Hermans, J. Modeling protein-small molecule interactions Structure and thermodynamics of noble gases binding in a cavity in mutant phage T4 lysozyme L99A, J. Mol. Biol. 2000, 302, 979-89. 

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