PDBsum entry 3hdf

Hydrolase

PDB id: 3hdf

Contents

Protein chains

133 a.a. *

Ligands

NO3 ×10

GOL ×6

Waters ×310

* Residue conservation analysis

PDB id:

3hdf

Name:

Hydrolase

Title:

Crystal structure of truncated endolysin r21 from phage 21

Structure:

Lysozyme. Chain: a, b. Fragment: unp residues 27-165. Synonym: lysis protein, muramidase, endolysin. Engineered: yes

Source:

Enterobacteria phage p21. Bacteriophage 21. Organism_taxid: 10711. Gene: r. Expressed in: escherichia coli. Expression_system_taxid: 562.

Resolution:

1.70Å R-factor: 0.200 R-free: 0.231

Authors:

Q.Sun,J.C.Sacchettini

Key ref:

Q.Sun et al. (2009). Regulation of a muralytic enzyme by dynamic membrane topology. Nat Struct Biol, 16, 1192-1194. PubMed id: 19881499 DOI: 10.1038/nsmb.1681

Date:

07-May-09 Release date: 03-Nov-09

Protein chains

P27359  (ENLYS_BPP21) -  SAR-endolysin from Enterobacteria phage P21

Seq: 165 a.a.

Struc: 133 a.a.*

* PDB and UniProt seqs differ at 2 residue positions (black crosses)

Enzyme reactions

• Enzyme class: E.C.3.2.1.17 - lysozyme.
• Reaction: Hydrolysis of the 1,4-beta-linkages between N-acetyl-D-glucosamine and N-acetylmuramic acid in peptidoglycan heteropolymers of the prokaryotes cell walls.

DOI no: 10.1038/nsmb.1681

Nat Struct Biol 16:1192-1194 (2009)

PubMed id: 19881499

Regulation of a muralytic enzyme by dynamic membrane topology.

Q.Sun, G.F.Kuty, A.Arockiasamy, M.Xu, R.Young, J.C.Sacchettini.

ABSTRACT

R(21), the lysozyme of coliphage 21, has an N-terminal signal-anchor-release (SAR) domain that directs its secretion in a membrane-tethered, inactive form and then its release and activation in the periplasm. Both genetic and crystallographic studies show that the SAR domain, once extracted from the bilayer, refolds into the body of the enzyme and effects muralytic activation by repositioning one residue of the canonical lysozyme catalytic triad.

Selected figure(s)

Figure 1.
(a) N-terminal sequences of R^21, Lyz^P1 and T4 E. The N-terminal domains of R^21, Lyz^P1 and T4 E are shown aligned by their Glu-8aa-Asp/Cys-5aa-Thr catalytic triad (blue and asterisks). SAR domains are boxed in orange. Leucine substitutions made in R^21 are shown above Gly14 and Gly15. The PelB signal sequence (PelB[ss]) and the artificial transmembrane domain (TMD[art]; yellow) are shown with arrows indicating points of fusion in the chimeric constructs. Residues involved in positioning the catalytic glutamate are highlighted by a light blue box in Lyz^P1 and T4 E. (b) Lysis profiles. ( ) pZE-luc, ( circle ) pZE-R^21[G14,15L], ( ) pZE-luc + 1 mM CHCl[3] at 80 min after induction, ( square ) pZE-R^21[G14,15L] + 1 mM CHCl[3] at 80 min after induction, ( ) pZE-R^21, ( diamond )Lyz^P1[1–26] R^21[27–165], ( ) pZE-pelB[ss] R^21[27–165], ( triangle ) pZE-TMD[art] R^21. A[550], absorbance at 550 nm. (c) Localization and processing of R^21 and derivatives after expression of the indicated chimeras. In top panel, lanes 1, 2 and 3 represent the total (T), periplasm (P) and spheroplast (S) fractions, respectively. Below, lanes 1, 2 and 3 represent the total (T), soluble (S) and membrane (M) fractions, respectively. (d) Morphologies of cells expressing the indicated pZE-R^21 plasmids at 100 min after induction. Scale bars (throughout) are 5 m.

Figure 2.
(a) Topological and conformational dynamics of R^21 activation. Crystal structures of ^iR^21 (left) and ^aR^21 (right) are represented in cartoon format, with -helix in cyan, -strand in magenta and coil in salmon; the SAR domain is shown in orange and is depicted as a membrane-spanning helix in ^iR^21. The catalytic triads are represented in stick-and-ball form, with disulfide linkages in yellow stick. (b) Alignment of the catalytic loop regions of ^iR^21 (green) and ^aR^21 (salmon). Except for Glu35, which shows a 10-Å C displacement (dashed line), most of the catalytic loop (Ser38–Thr67) of ^aR^21 can be superimposed on the same region of ^iR^21 (r.m.s. difference = 0.4 Å for 209 atoms). (c) Alignment of the C-terminal domains of ^iR^21 and ^aR^21, colored as in b. Beginning at Glu96, the backbone r.m.s. difference between the two structures is 4.0 Å. Helix 8 in ^iR^21 tilts 30°, turns and unwinds compared to ^aR^21, thereby turning Arg152 away from Glu35. (d) Polarity switching at the interface for the SAR domain. The calculated electrostatic surface (positive = blue; negative = red) contacting the helices of the extracted SAR domain for ^aR^21 is shown at right; the corresponding surface is ^iR^21 is shown at left, with the SAR helices superimposed as an orange backbone ribbon trace.

The above figures are reprinted by permission from Macmillan Publishers Ltd: Nat Struct Biol (2009, 16, 1192-1194) copyright 2009.