**5.2 Aminotransferase A**

*Aminotransferase A* (ATA, EC 2.6.1) is a PLP-dependant enzyme that catalyses the conversion of NAAKP and glutamate to N-acetyl-(2*S*)-2,6,-diaminopimelate (NADAP) and αketoglutarate (Ledwidge & Blanchard., 1999; Peterkofsky & Gilvarg., 1961) (Fig. 1). It has been speculated that the ATA reaction mechanism resembles that of NSDAP-AT (Section 4.2) (Ledwidge & Blanchard., 1999; Peterkofsky & Gilvarg., 1961). Crude cell extracts from *B. megaterium* were found to contain active ATA (Chatterjee & White., 1982), with ATA activity identified by monitoring enzyme activity in the reverse direction utilising an acid ninhydrin assay (Chatterjee & White., 1982; Sundharadas & Gilvarg., 1967).

#### **5.3 N-acetyldiaminopimelate deacetylase**

*N-acetyldiaminopimelate deacetylase* (NAD-DAC, EC 3.5.1.47) catalyses the hydrolysis of NADAP to form DAP and acetate (Fig. 1). NAD-DAC was first identified from studies involving the isolation of a *B. megaterium* DAP auxotroph (Saleh & White., 1979; Sundharadas & Gilvarg., 1967). The mutant strain possesses a non-functional form of NAD-

The catalytic domain incorporates a negatively charged active site cleft, containing two zinc ions. One zinc ion is coordinated by the imidazole group and sidechain oxygens of His67 and Glu163, respectively, whilst another zinc ion is coordinated in a similar manner by His349 and Glu135. The zinc ions are bridged together by interaction with Asp100 and a

The availability of a structural model has resulted in a proposed mechanism for hydrolysis of NSDAP by SDAP-DS (Born et al., 1998; Nocek et al., 2010). It is hypothesised that NSDAP adopts an extended conformation when bound to the active site of the enzyme. The NSDAP amide carbonyl coordinates to an active site Zn2+ ion and becomes avaliable for nucleophilic attack. This binding event displaces a bridging water molecule, resulting in its hydrolysis by Glu134 and the generation of a zinc bound nucleophilic hydroxide. The hydroxide then attacks the target carbonyl carbon to form a η-1-μ-transition-state complex, which then

The DAP isomers *L,L*-DAP and *D,L*-DAP are competitive inhibitors of *H. influenzae* SDAP-DS, exhibiting *K*i values of 8 and 12 mM, respectively (Born et al., 1998). Studies employing Zn K-edge EXAFS suggest that the *H. influenzae* SDAP-DS inhibitor, 5-mercaptopentanoic

*Tetrahydrodipicolinate N-acetyltransferase* (THDP-NAT, EC 2.3.1.89) is an acetyl-coenzyme A (ACoA) dependant enzyme that catalyses the conversion of cyclic THDP to acyclic N-acetyl- (*S*)-2-amino-6-ketopimelate (NAAKP) (Chatterjee & White., 1982) (Fig. 1). The transferred acyl group maintains the linear conformation of the product and exposes the 6-keto group for subsequent transamination (Beaman et al., 2002). Crude cell extracts from *B. megaterium* were found to contain active THDP-NAT (Chatterjee & White., 1982). THDP-NAT enzymes

*Aminotransferase A* (ATA, EC 2.6.1) is a PLP-dependant enzyme that catalyses the conversion of NAAKP and glutamate to N-acetyl-(2*S*)-2,6,-diaminopimelate (NADAP) and αketoglutarate (Ledwidge & Blanchard., 1999; Peterkofsky & Gilvarg., 1961) (Fig. 1). It has been speculated that the ATA reaction mechanism resembles that of NSDAP-AT (Section 4.2) (Ledwidge & Blanchard., 1999; Peterkofsky & Gilvarg., 1961). Crude cell extracts from *B. megaterium* were found to contain active ATA (Chatterjee & White., 1982), with ATA activity identified by monitoring enzyme activity in the reverse direction utilising an acid ninhydrin

*N-acetyldiaminopimelate deacetylase* (NAD-DAC, EC 3.5.1.47) catalyses the hydrolysis of NADAP to form DAP and acetate (Fig. 1). NAD-DAC was first identified from studies involving the isolation of a *B. megaterium* DAP auxotroph (Saleh & White., 1979; Sundharadas & Gilvarg., 1967). The mutant strain possesses a non-functional form of NAD-

acid, may exert its effect through binding to active site Zn2+ ions (Cosper et al., 2003).

are thought to be largely confined to *Bacillus sp*. (Weinberger & Gilvarg., 1970).

assay (Chatterjee & White., 1982; Sundharadas & Gilvarg., 1967).

**5.3 N-acetyldiaminopimelate deacetylase** 

water/hydroxide.

resolves to release DAP and succinate.

**5.1 Tetrahydrodipicolinate N-acetyltransferase** 

**5. Acetylase pathway** 

**5.2 Aminotransferase A** 

DAC and consequently accumulates NADAP. Early studies of this enzyme centred on Gram-positive species, with NAD-DAC activity identified by utilising an acid ninhydrin assay to detect NADAP formation in crude cell extracts (Chatterjee & White., 1982; Weinberger & Gilvarg., 1970). The distribution of NAD-DAC has since been investigated in large number of Gram-negative and Gram-positive bacteria. Interestingly, the enzyme appears to be restricted to *Bacillus sp*. (Weinberger & Gilvarg., 1970).
