Good-chop, bad-chop? Defining the activities of calpain-like cysteine proteases from oral Actinomyces bacteria

Dr Watt, Rory Munro   (Principal Investigator (PI))

Professor Ngai Sai Ming   (Co-Investigator)

Professor Bartlam Mark Gerrard   (Co-Investigator)

42

2018-09-01

2022-02-28

966838

Good-chop, bad-chop? Defining the activities of calpain-like cysteine proteases from oral Actinomyces bacteria

bacterial infection, caries, cysteine protease, oral microbiome, protein biochemistry

Dentistry,Microbiology

Biology and Medicine (M)

17103318

General Research Fund (GRF)

2018

Completed

1 Dissect the biochemical activities of actinopain peptidases from three distinct evolutionary lineages. Screen their peptide substrate specificities, and determine their abilities to cleave proteins/peptides of biological relevance. Map their autolytic processing patterns into catalytically-active/inactive mature forms. 2 Use protein X-ray crystallography and modelling-based approaches to elucidate structure-activity relationships within actinopain- and bacterial calpain-family proteases. Create selected point-mutated actinopain proteins to test/validate structural and mechanistic predictions. 3 Utilize activity-based chemical probes that target calpain-family cysteine proteases to label actinopain peptidases in Actinomyces cells, to characterize protein expression patterns during cultivation under different biologically-relevant conditions. Compare and contrast results with data obtained from analogous transcriptional analyses. 4 Create actinopain gene-deletion mutants in selected Actinomyces species, to analyze how actinopain peptidase expression affects cellular growth and phenotypic properties; inter-species coaggregation activities; biofilm-forming capabilities, and infectivity.