Streptococcus pneumoniae Aspartate Semialdehyde-Dependent Dehydrogenase Initiates Polyamine Biosynthesis with Agmatine

Streptococcus pneumoniae is a major bacterial pathogen and the leading cause of community-acquired pneumonia. Polyamines, particularly spermidine, play a key role in pneumococcal virulence. However, spermidine biosynthesis in S. pneumoniae remains incompletely characterized. We predicted that the S. pneumoniae aspartate semialdehyde dehydrogenase (also known as carboxyspermidine dehydrogenase or CASDH) would use agmatine as its polyamine substrate rather than putrescine, as was previously expected. S. pneumoniae CASDH substrate specificity was evaluated using steady-state kinetic NADPH oxidation assays in the presence of agmatine and putrescine (as well as diaminopropane, cadaverine, N-carbamoylputrescine, and spermidine). CASDH displayed the greatest catalytic efficiency toward agmatine with a kcat/KM of 1.0 × 106 M−1s−1, approximately 3200-fold greater than putrescine and indicating that the S. pneumoniae enzyme is a carboxyaminopropylagmatine dehydrogenase or CAPADH and not a carboxyspermidine dehydrogenase or CASDH. To confirm the residues involved in agmatine specificity we solved a 1.19 Å X-ray crystal structure of CAPADH with agmatine and NADPH bound providing evidence that a conserved Asn-Glu-Phe-Glu motif coordinates agmatine. These results require a reconsideration of the biosynthetic steps leading to spermidine production in S. pneumoniae and provide the first structural evidence for substrate specificity in the aspartate semialdehyde dehydrogenase enzyme class.