21 Vertebrate Lysozymes

Imoto, Taiji; Johnson, L.N.; North, A. C. T.; Phillips, David; Rupley, John A.

doi:10.1016/s1874-6047(08)60465-5

Cited by 494 publications

(410 citation statements)

References 517 publications

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“…Previous studies on model proteins reported that partial modification of lysines or arginines is compatible with the maintenance of native-like structures (Imoto et al, 1972;Glocker et al, 1994;Przybylski & Glocker, 1996). These findings are consistent with the observation that charged residues are usually exposcd on the protein surface and mainly involved in protein-solvent interactions.…”

Section: Discussionmentioning

confidence: 98%

Surface topology of Minibody by selective chemical modifications and mass spectrometry

et al. 1997

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The surface topology of the Minibody, a small de novo-designed P-protein, has been probed by a strategy that combines selective chemical modification with a variety of reagents and mass spectrometric analysis of the modified fragments. Under appropriate conditions, the susceptibility of individual residues primarily depends on their surface accessibility so that their relative reactivities can be correlated with their position in the tertiary structure of the protein. Moreover, this approach provides information on interacting residues, since intramolecular interactions might greatly affect the reactivity of individual side chains by altering their pKa values.The results of this study indicate that, while overall the Minibody model is correct, the P-sheet formed by the N-and C-terminal segments is most likely distorted. This is also in agreement with previous results that were obtained using a similar approach where mass spectrometry was used to identify Minibody fragments from limited proteolysis (Zappacosta F, Pessi A, Bianchi E, Venturini s, Sollazzo M, Tramontano A, Marino G, Pucci P. 1996. Probing the tertiary structure of proteins by limited proteolysis and mass spectrometry: The case of Minibody. Protein Sci 5902-813). The chemical modification approach, in combination with limited proteolysis procedures, can provide useful, albeit partial, structural information to complement simulation techniques. This is especially valuable when, as in the Minibody case, an NMR and/or X-ray structure cannot be obtained due to insufficient solubility of the molecule.

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Section: Discussionmentioning

confidence: 98%

Surface topology of Minibody by selective chemical modifications and mass spectrometry

et al. 1997

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“…Alternatively, HEWL binding may occur at any site along the glycan backbone, but with weaker affinities at sites of 0-acetylation. Analysis of the X-ray crystallographic structures of HEWL-inhibitor complexes [26] indicate that contacts are made between the enzyme and C6 hydroxyl groups of muramyl residues. Thus, it is plausible that acetyl modification at these positions either precludes or constrains HEWL binding.…”

Section: Discussionmentioning

confidence: 99%

Dependence of lysozyme‐catalysed solubilization of Proteus mirabilis peptidoglycan on the extent of O‐acetylation

Dupont

Clarke

1991

European Journal of Biochemistry

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The degree of peptidoglycan 0-acetylation in 14 strains of Proteus mirabilis has been accurately determined by a procedure which employs the quantitation of mild-base-released acetic acid by HPLC, and the estimation of peptidoglycan concentration by cation-exchange amino acid analysis. The /3-~-N,6-O-diacetylmuramyl content of all isolated and purified peptidoglycans was ranged 20-52.8%, relative to the total muramic acid concentration. Each of the 0-acetylated peptidoglycans was found to be resistant to solubilization by both human and hen eggwhite lysozymes and for hen egg-white lysozyme, the extent of this resistance was dependent upon the degree of 0-acetylation. The steady-state parameters, K, and V, for the hen-egg-white-lysozyme-catalysed solubilization of various peptidoglycan preparations were determined at pH 6.61 and 25°C. Values of K , for the different peptidoglycan samples were found to increase with increasing 0-acetylation, whereas with V no such relationship appeared to exist. An increase in the overall change in the standard Gibbs free energy of activation [d(dG*)], a consequence of increasing 0-acetylation, was observed, and is shown to result from the weaker affinity of the enzyme for the modified substrates.Peptidoglycan is a heteropolymer of distinct composition and structure, associated uniquely with eubacterial cell walls. Proteus mirabilis peptidoglycan is comprised of equimolar amounts of GlcNAc, N-acetylmuramic acid (MurNAc), Lalanine, D-glutamic acid, L-meso-diaminopimelic acid and Dalanine. The cross-linking of neighbouring peptides is direct (chemotype Aly) and involves the meso-diaminopimelic acid and D-alanyl residues. The extent of cross-linking (33%) is typical of most Gram-negative bacteria [l]. Thus, the peptidoglycan of P. mirabilis is characteristic of the Enterobacteriaceae but for one notable and significant difference, the presence of extensive 0-acetylation [2]. Peptidoglycan 0-acetylation occurs at the 6-hydroxyl group of MurNAc residues producing the corresponding N,O-diacetylmuramyl derivatives, and this modification confers resistance to the hydrolytic action of many muramidases, including hen egg-white lysozyme (HEWL) and human lysozymes [3 -61.The phenomenon of peptidoglycan 0-acetylation was first observed independently by two different groups of researchers over 30 years ago [7, 81, and its role in conferring lysozyme resistance was demonstrated soon after [6]. Since then, 0-acetylation of peptidoglycan has been observed in a diverse group of bacteria which includes some important pathogens, both Gram positive (e.g. Staphylococcus aureus [9, 101) and Gram negative (e. g. Neisseria gonorrhoeae [3] and P. mirabilis strains of N . gonorrhoeae and characterized by 34-52% 0-acetylation, was shown to be resistant to hydrolysis by HEWL to the extent of 34-59% [12]. On the other hand, O-acetyldeficient peptidoglycan, isolated from N . gonorrhoea RD5 was completely solubilized and degraded to low-molecular-mass fragments. In the study, resistance was defined as the percen...

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“…Recently, the structure of turkey egg-white lysozyme was published (7), and it was shown that the E and F sites of this enzyme were not blocked in the crystal (7), as was the case for the crystal of hen egg-white lysozyme (5,6) used in this study. Hopefully, therefore, it will be possible to obtain crystal structures of saccharides bound to the E and F regions so that experimental structural information concerning the complex will be available.…”

Section: Methodsmentioning

confidence: 67%

“…We are currently investigating this possibility. In a proposed structure of the lysozyme-hexamer complex based on model building (5), the fifth residue (in site E) from the nonreducing end had Unfortunately, there are few experimental data with which to compare our calculated lowest-energy binding position for the fifth residue and the one proposed from model building (5). There are experimental data (6) (5).…”

Section: Methodsmentioning

confidence: 99%

“…Now that we have carried out an investigation of possible binding regions for oligosaccharides at the active site of the rigid enzyme, it is of interest to compare the relative affinities of each site available to the oligomers. Table 2 presents the relative affinities (Ecom) that were obtained from a minimization of the conformational energy of the monomer (GlcNAc) in each binding region, along with free energies of binding determined from experimental data of Imoto et al (5). The calculated data gave only approximate binding energies for oligomer residues because the monomers bound in slightly different orientations from those of the corresponding residues of oligomers, i.e., there were no constraints of an oligomer chain on the binding of a monomer.…”

Section: Methodsmentioning

confidence: 99%

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Structures of enzyme-substrate complexes of lysozyme

Pincus

Zimmerman

Scheraga

1977

Proc. Natl. Acad. Sci. U.S.A.

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Conformational energy calculations were used to determine the binding structures of two oligosaccharides (GIcNAc),, in which n = 5 and 6, in the rigid active site of lysozyme (mucopeptide N-acetylmuramoylhydrolase, EC 3.2.1.17) Starting with the lowest energy binding structures of (GlcNAc)4 as determined in a previous publication, we added a fifth GlcNAc residue to this tetramer in three different conformations, corresponding to the left-handed and right-handed helical structures and an intermediate structure, and the energy of each complex was minimized. The most stable binding conformation of the fifth residue of the pentamer was closest to the lefthanded helical one. During energy minimization, the fourth residue of the pentamer moved from its initial position near the surface of the active site farther into the active site cleft at binding site D. Binding structures of (GIcNAc)o were then examined by addition of a residue to the lowest energy structure of (GlcNAc).5, and it was found that the sixth residue of the hexamer binds in a conformation again close to the left-handed helical one. Stable binding regions of the rigid active site for the fifth and sixth residues were found to be near arginyl 45 and asparaginyl 46, on the opposite side of the active site cleft from arginyl 114. When the calculated structure of the lysozyme-(GlcNAc)4 complex (used here as the starting structure for addition of the fifth and sixth residues) is compared with recent experimental data, it is found that the calculated structure is a reasonable one. Of all binding regions available to the saccharide residues, the C site binds GlcNAc with the lowest energy, in agreement with experiments.In a previous publication (1), we presented an analysis of the manner in which oligosaccharides (substrates and inhibitors) containing up to four units of GlcNAc bound to the rigid active site of the enzyme lysozyme (mucopeptide N-acetylmuramoylhydrolase, EC 3.2.1.17). In that analysis, we centered our attention on the first four contiguous binding regions of the active site (i.e., sites A through D). We now point out that the lowest-energy structure of the lysozyme-(GlcNAc)4 (lysozyme-tetra-N-acetyl-D-glucosamine) complex determined earlier (1) is in agreement with experimental data published subsequently (2). We also show here that successive GlcNAc residues can be added to the tetramer structure (proposed in ref. 1) to give unique low-energy structures for the lysozyme-(GlcNAc)5 and lysozyme-(GlcNAc)6 complexes.It must be emphasized that in this paper, as in the earlier one (1), the active site of the enzyme is assumed to be rigid throughout the calculations; only the atoms of the substrate are allowed to move. The purpose here is to determine whether the rigid native enzyme possesses the structural information that allows for the recognition of the substrate. Because we have not minimized the energy of the isolated enzyme, the results presented here are based on the reported x-ray coordinates and could be altered if the x-ray coordi...

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21 Vertebrate Lysozymes

Cited by 494 publications

References 517 publications

Surface topology of Minibody by selective chemical modifications and mass spectrometry

Surface topology of Minibody by selective chemical modifications and mass spectrometry

Dependence of lysozyme‐catalysed solubilization of Proteus mirabilis peptidoglycan on the extent of O‐acetylation

Structures of enzyme-substrate complexes of lysozyme

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