The autolytic activity of the recombinant amidase ofStaphylococcus saprophyticusis inhibited by its own recombinant GW repeats

Abstract

The Aas (autolysin/adhesin of Staphylococcus saprophyticus) is a multifunctional surface protein containing two enzymatic domains an N-acetyl-muramyl-L-alanine amidase, an endo-L-N-acetyl-D-glucosaminidase, and two different regions of repetitive sequences, an N-terminal and a C-terminal repetitive domain. The C-terminal repetitive domain is built up by the repeats R1, R2 and R3, which interconnect the putative active centers of the amidase and glucosaminidase. To investigate the influence of the C-terminal repeats and the N-terminal repeats on the amidase activity, the repetitive domains and fragments of them were cloned and expressed in Escherichia coli. The influence of the different fragments on the activity of the recombinant amidase of the Aas, consisting of the active center of the enzyme and repeat R1, was investigated in a turbidimetric microassay. The different fragments derived from the C-terminal repeats inhibited the amidase activity, while the N-terminal repeats did not influence the activity of the enzyme. The inhibiting activity increased with the number of GW repeats the recombinant fragment contained. Thus we conclude, that the C-terminal GW repeats and not the N-terminal repeats are necessary for the cell wall targeting and the autolytic function of the amidase.

Molecular properties of the putative autolysin AtlWM encoded by Staphylococcus warneri M: Mutational and biochemical analyses of the amidase and glucosaminidase domains

“…More recently, Biswas et al (2006) purified three amidase domains of the S. epidermidis AtlE: amiE with no repeat regions, amiE-R1-R2, and the three repeats (R1-R2-R3); they speculated that AmiE indeed cleaves the amide bond between N-acetyl muramic acid and L-alanine, and the repeat sequences R1-R2-R3 probably have a higher affinity to premature peptidoglycan structures. Hell et al (2003) also reported similar results that repeat sequences from autolysin/adhesin protein Aas of S. saprophyticus function in the cell wall targeting.…”

Identification of IspC, an 86-Kilodalton Protein Target of Humoral Immune Response to Infection with Listeria monocytogenes Serotype 4b, as a Novel Surface Autolysin

“…This suggests that the IspC cell wall binding domain may be of importance in modulating the activity of the N-terminal enzyme domain. The importance of the C-terminal GW repeats for the autolytic function of an enzyme has been described for Atl in S. aureus (2) and Aas in Staphylococcus saprophyticus (21). Similarly, targeting of IspC to the cell wall through its C-terminal region containing GW modules may be necessary for the autolytic function.…”

The Multifunctional Staphylococcus aureus Autolysin Aaa Mediates Adherence to Immobilized Fibrinogen and Fibronectin

“…This domain consists of GW-containing repeats. Such GW repeats are also present within the repeat regions of the homologous autolysins Atl from S. aureus, AtlE from S. epidermidis, and Aas from S. saprophyticus (23,42). Like these major staphylococcal autolysins, Aaa contains three repetitive sequences, but they are not homologous to the GW repeats.…”