Structural studies examining the substrate specificity profiles of PC-PLCBc protein variants

Aaron P. Benfield, Nina M. Goodey, Lauren T. Phillips, Stephen F. Martin

Research output: Contribution to journalArticle

16 Citations (Scopus)

Abstract

The phosphatidylcholine preferring phospholipase C from Bacillus cereus (PC-PLCBc) catalyzes the hydrolysis of phospholipids in the following order of preference: phosphatidylcholine (PC) > phosphatidylethanolamine (PE) > phosphatidylserine (PS). In previous work, mutagenic, kinetic, and crystallographic experiments suggested that varying the amino acids at the 4th, 56th, and 66th positions had a significant influence upon the substrate specificity profile of PC-PLCBc. Here, we report the crystal structures of the native form of several PC-PLCBc variants that exhibited altered substrate specificities for PC, PE, and PS at maximum resolutions of 1.90-2.05 Å. Comparing the structures of these variants to the structure of the wild-type enzyme reveals only minor differences with respect to the number and location of active site water molecules and the side chain conformations of residues at the 4th and 56th positions. These results suggest that subtle changes in steric and electronic properties in the substrate binding site of PC-PLCBc are responsible for the significant changes in substrate selectivity.

Original languageEnglish
Pages (from-to)41-47
Number of pages7
JournalArchives of Biochemistry and Biophysics
Volume460
Issue number1
DOIs
StatePublished - 1 Apr 2007

Fingerprint

Bacillus cereus
Type C Phospholipases
Substrate Specificity
Phosphatidylcholines
Substrates
Proteins
Phosphatidylserines
Electronic properties
Conformations
Hydrolysis
Catalytic Domain
Phospholipids
Crystal structure
Binding Sites
Amino Acids
Molecules
Kinetics
Water
Enzymes

Keywords

  • Bacillus cereus
  • Crystallography
  • Phosphatidylcholine
  • Phospholipase C
  • Substrate specificity

Cite this

Benfield, Aaron P. ; Goodey, Nina M. ; Phillips, Lauren T. ; Martin, Stephen F. / Structural studies examining the substrate specificity profiles of PC-PLCBc protein variants. In: Archives of Biochemistry and Biophysics. 2007 ; Vol. 460, No. 1. pp. 41-47.
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Structural studies examining the substrate specificity profiles of PC-PLCBc protein variants. / Benfield, Aaron P.; Goodey, Nina M.; Phillips, Lauren T.; Martin, Stephen F.

In: Archives of Biochemistry and Biophysics, Vol. 460, No. 1, 01.04.2007, p. 41-47.

Research output: Contribution to journalArticle

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AB - The phosphatidylcholine preferring phospholipase C from Bacillus cereus (PC-PLCBc) catalyzes the hydrolysis of phospholipids in the following order of preference: phosphatidylcholine (PC) > phosphatidylethanolamine (PE) > phosphatidylserine (PS). In previous work, mutagenic, kinetic, and crystallographic experiments suggested that varying the amino acids at the 4th, 56th, and 66th positions had a significant influence upon the substrate specificity profile of PC-PLCBc. Here, we report the crystal structures of the native form of several PC-PLCBc variants that exhibited altered substrate specificities for PC, PE, and PS at maximum resolutions of 1.90-2.05 Å. Comparing the structures of these variants to the structure of the wild-type enzyme reveals only minor differences with respect to the number and location of active site water molecules and the side chain conformations of residues at the 4th and 56th positions. These results suggest that subtle changes in steric and electronic properties in the substrate binding site of PC-PLCBc are responsible for the significant changes in substrate selectivity.

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