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Abstract

Bacteria frequently adapt to high osmolarity surroundings through the
accumulation of compatible solutes. Ectoine is a prominent member of
these types of stress protectants and is produced via an evolutionarily
conserved biosynthetic pathway beginning with the L-2,4-diaminobutyrate
(DAB) transaminase (TA) EctB. Here, we studied EctB from the
thermo-tolerant Gram-positive bacterium Paenibacillus lautus (Pl) and
show that this tetrameric enzyme is highly tolerant to salt, pH, and
temperature. During ectoine biosynthesis, EctB converts L-glutamate and
L-aspartate-beta-semialdehyde into 2-oxoglutarate and DAB, but it also
catalyzes the reverse reaction. Our analysis unravels that EctB enzymes
are mechanistically identical to the PLP-dependent gamma-aminobutyrate
TAs (GABA-TAs) and only differ with respect to substrate binding.
Inspection of the genomic context of the ectB gene in P. lautus
identifies an unusual arrangement of juxtapositioned genes for ectoine
biosynthesis and import via an Ehu-type binding-protein-dependent ABC
transporter. This operon-like structure suggests the operation of a
highly coordinated system for ectoine synthesis and import to maintain
physiologically adequate cellular ectoine pools under osmotic stress
conditions in a resource-efficient manner. Taken together, our study
provides an in-depth mechanistic and physiological description of EctB,
the first enzyme of the ectoine biosynthetic pathway.