Taxonomic group: bacteria / Proteobacteria
(Phylum: Proteobacteria)
NCBI PubMed ID: 36346293Publication DOI: 10.1002/pro.4502Journal NLM ID: 9211750Publisher: Woodbury, NY: Cold Spring Harbor Laboratory Press for the Protein Society
Correspondence: Hazel_Holden
biochem.wisc.edu
Institutions: Department of Biochemistry, University of Wisconsin, Madison, WI, USA
Psychrobacter cryohalolentis K5T is a Gram-negative bacterium first isolated from Siberian permafrost in 2006. It has a complex O-antigen containing l-rhamnose, d-galactose, two diacetamido-sugars, and one triacetamido-sugar. The biosynthetic pathway for one of the diacetamido-sugars, namely 2,3-diacetamido-2,3-dideoxy-d-glucuronic acid, is presently unknown. Utilizing the published genome sequence of P. cryohalolentis K5T , we hypothesized that the genes designated Pcryo_0613, Pcryo_0614, Pcryo_0616, and Pcryo_0615 encode for a uridine dinucleotide (UDP)-N-acetyl-d-glucosamine 6-dehydrogenase, an nicotinamide adenine dinucleotide (oxidized) (NAD+ )-dependent dehydrogenase, a pyridoxal 5'-phosphate (PLP)-dependent aminotransferase, and an N-acetyltransferase, respectively, activities of which would be required for the biosynthesis of this unusual carbohydrate. Here we present the cloning, overexpression, and purification of these hypothetical proteins. Kinetic data on the enzymes encoded by Pcryo_0613, Pcryo_0614, and Pcryo_0615 confirmed their postulated biochemical activities. In addition, the high-resolution X-ray structures of both the internal and external aldimine forms of the aminotransferase were determined to 1.25 and 1.0 Å, respectively. Finally, the three-dimensional architecture of the N-acetyltransferase in complex with its substrate and coenzyme A was solved to 1.8 Å resolution. Strikingly, the N-acetyltransferase was shown to adopt a new motif for UDP-sugar binding. The data presented herein provide additional insight into sugar biosynthesis in Gram-negative bacteria.
O-antigen, carbohydrate, 2, 3-diacetamido-2, 3-dideoxy-D-glucuronic acid, aminotransferase, N-acetyltransferase, enzyme structure, 3-dideoyhexose, Psychrobacter cryohalolentis K5T
Structure type: polymer chemical repeating unit
Location inside paper: scheme 1
Compound class: O-polysaccharide, O-antigen
Contained glycoepitopes: IEDB_133754,IEDB_136105,IEDB_136906,IEDB_137472,IEDB_141794,IEDB_142345,IEDB_151528,IEDB_190606,IEDB_225177,IEDB_885823,SB_7
Methods: X-ray, kinetics assays, biochemical methods, crystallization, biochemical synthesis
3D data: 3D structures of Pcryo_0616, Pcryo_0615
NCBI Taxonomy refs (TaxIDs): 330922
Show glycosyltransferases
There is only one chemically distinct structure:
Found 
report error