ISOLATION OF THE PUTP GENE OF CORYNEBACTERIUM-GLUTAMICUM AND CHARACTERIZATION OF A LOW-AFFINITY UPTAKE SYSTEM FOR COMPATIBLE SOLUTES

Corynebacterium glutamicum accumulates the compatible solutes proline, glycine betaine, and ectoine under conditions of high osmolality. Upt ake of proline is mediated by both a high-affinity and a low-affinity secondary transport system. The low-affinity uptake system also accept s glycine betaine and ectoine as substrates. In the present study, the gene encoding the high-affinity proline uptake system PutP was isolat ed by heterologous complementation of Escherichia coli mutant strain W G389, which lacks the transport systems BetT, PutP, ProP, and ProU and is unable to synthesize proline and glycine betaine. This gene (putP) encodes a protein of 524 amino acids that shares identity with the pr oline transport systems PutP of E. coli, Staphylococcus aureus, Salmon ella typhimurium, Haemophilus influenzae, and Klebsiella pneumoniae. F unctional studies of PutP synthesized in E. coli mutant strain MKH13, which also lacks the transport systems for compatible solutes and is u nable to synthesize glycine betaine, revealed that this carrier system is not regulated by the external osmolality on the level of activity. K-m values of 7.6 mM for proline and 1.3 mM for sodium as cotransport ed ion were determined. Deletion of the putP gene allowed the function al characterization of another proline uptake system with low affinity .