Identification and functional analysis of fructosyl amino acid-binding protein from Gram-positive bacterium Arthrobacter sp.
OBJECTIVE
Fructosyl amino acid-binding protein (FABP) is a substrate-binding protein (SBP), which recognizes fructosyl amino acids (FAs) as its ligands. Although FABP has been shown as a molecular recognition tool of biosensing for glycated proteins, the availability of FABP is still limited and no FABP was reported from Gram-positive bacteria. In this study, a novel FABP from Gram-positive bacteria, Arthrobacter spp., was reported.


RESULTS
BLAST analysis revealed that FABP homologues exist in some of Arthrobacter species genomes. An FABP homologue cloned from Arthrobacter sp. FV1-1, FvcA, contained a putative lipoprotein signal sequence, suggesting that it is a lipoprotein anchored to the bacterial cytoplasmic membrane, which is a typical characteristic for SBPs from Gram-positive bacteria. In contrast, FvcA also exhibits high amino acid sequence similarity to a known Gram-negative bacterial FABP, which exists as a free periplasmic protein. FvcA, without the N-terminal anchoring region, was then recombinantly produced as soluble protein and was found to exhibit Nα-FA-specific binding activity by intrinsic fluorescent measurement.


CONCLUSIONS
This study identified a novel FABP from a Gram-positive bacterium, Arthrobacter sp., which exhibited Nα-FA-specific binding ability. This is the first report concerning an FABP from a Gram-positive bacterium, suggesting that FABP-dependent FA catabolism system is also present in Gram-positive bacteria.


CONCLUSIONS
The novel FABP exhibits the ability to specifically bind to Nα-FA with a high affinity. This selectivity is beneficial for applying FABP in HbA1c sensing. The successful preparation of water-soluble, functionally expressed Gram-negative bacterial FABP may make way for future applications for a variety of SBPs from Gram-positive bacteria employing the same expression strategy. The results obtained here enhance our understanding of bacterial FA catabolism and contribute to the improved development of FABP as Nα-FA-sensing molecules.
Graduate School of Bionics, Computer and Media Sciences, Tokyo University of Technology, Hachioji, Japan.
A
A
Sakaguchi-Mikami
S
S
Ferri
S
S
Katayama
W
W
Tsugawa
K
K
Sode
D000595:Amino Acid Sequence; D001173:Arthrobacter / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D005227:Fatty Acids / Q000378:metabolism; D019270:Fructosamine / Q000378:metabolism*; D008024:Ligands; D008074:Lipoproteins / Q000235:genetics / Q000378:metabolism; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D021382:Protein Sorting Signals; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
D001426:Bacterial Proteins; D005227:Fatty Acids; D008024:Ligands; D008074:Lipoproteins; D008565:Membrane Proteins; D021382:Protein Sorting Signals; D019270:Fructosamine