Kobus S

References (3)

Title : Crystal structures of a novel family IV esterase in free and substrate-bound form - Hoppner_2021_FEBS.J_288_3570
Author(s) : Hoppner A , Bollinger A , Kobus S , Thies S , Coscolin C , Ferrer M , Jaeger KE , Smits SHJ
Ref : Febs J , 288 :3570 , 2021
Abstract : Bacterial lipolytic enzymes of family IV are homologs of the mammalian hormone-sensitive lipases (HSL) and have been successfully used for various biotechnological applications. The broad substrate specificity and ability for enantio-, regio-, and stereoselective hydrolysis are remarkable features of enzymes from this class. Many crystal structures are available for esterases and lipases, but structures of enzyme-substrate or enzyme-inhibitor complexes are less frequent although important to understand the molecular basis of enzyme substrate interaction and to rationalize biochemical enzyme characteristics. Here, we report on the structures of a novel family IV esterase isolated from a metagenomic screen which shows a broad substrate specificity. We solved the crystal structures in the apo form and with a bound substrate analogue at 1.35 and 1.81 resolution, respectively. This enzyme named PtEst1 hydrolyzed more than 60 out 96 structurally different ester substrates thus being substrate promiscuous. Its broad substrate specificity is in accord with a large active site cavity, which is covered by an alpha-helical cap domain. The substrate analogue methyl 4-methylumbelliferyl hexylphosphonate was rapidly hydrolyzed by the enzyme leading to a complete inactivation caused by covalent binding of phosphinic acid to the catalytic serine. Interestingly, the alcohol leaving group 4-methylumbelliferone was found remaining in the active site cavity and additionally, a complete inhibitor molecule was found at the cap domain next to the entrance of the substrate tunnel. This unique situation allowed gaining valuable insights into the role of the cap domain for enzyme-substrate interaction of esterases belonging to family IV.
ESTHER : Hoppner_2021_FEBS.J_288_3570
PubMedSearch : Hoppner_2021_FEBS.J_288_3570
PubMedID: 33342083
Gene_locus related to this paper: pseth-a0a1m6y2k1

Title : A Novel Polyester Hydrolase From the Marine Bacterium Pseudomonas aestusnigri - Structural and Functional Insights - Bollinger_2020_Front.Microbiol_11_114
Author(s) : Bollinger A , Thies S , Knieps-Grunhagen E , Gertzen C , Kobus S , Hoppner A , Ferrer M , Gohlke H , Smits SHJ , Jaeger KE
Ref : Front Microbiol , 11 :114 , 2020
Abstract : Biodegradation of synthetic polymers, in particular polyethylene terephthalate (PET), is of great importance, since environmental pollution with PET and other plastics has become a severe global problem. Here, we report on the polyester degrading ability of a novel carboxylic ester hydrolase identified in the genome of the marine hydrocarbonoclastic bacterium Pseudomonas aestusnigri VGXO14T. The enzyme, designated PE-H, belongs to the type IIa family of PET hydrolytic enzymes as indicated by amino acid sequence homology. It was produced in Escherichia coli, purified and its crystal structure was solved at 1.09 A resolution representing the first structure of a type IIa PET hydrolytic enzyme. The structure shows a typical alpha/beta-hydrolase fold and high structural homology to known polyester hydrolases. PET hydrolysis was detected at 30C with amorphous PET film (PETa), but not with PET film from a commercial PET bottle (PETb). A rational mutagenesis study to improve the PET degrading potential of PE-H yielded variant PE-H (Y250S) which showed improved activity, ultimately also allowing the hydrolysis of PETb. The crystal structure of this variant solved at 1.35 A resolution allowed to rationalize the improvement of enzymatic activity. A PET oligomer binding model was proposed by molecular docking computations. Our results indicate a significant potential of the marine bacterium P. aestusnigri for PET degradation.
ESTHER : Bollinger_2020_Front.Microbiol_11_114
PubMedSearch : Bollinger_2020_Front.Microbiol_11_114
PubMedID: 32117139
Gene_locus related to this paper: 9psed-peh

Title : Structural basis for recognition and ring-cleavage of the Pseudomonas quinolone signal (PQS) by AqdC, a mycobacterial dioxygenase of the alpha\/beta-hydrolase fold family - Wullich_2019_J.Struct.Biol_207_287
Author(s) : Wullich SC , Kobus S , Wienhold M , Hennecke U , Smits SHJ , Fetzner S
Ref : J Struct Biol , 207 :287 , 2019
Abstract : The cofactor-less dioxygenase AqdC of Mycobacteroides abscessus catalyzes the cleavage and thus inactivation of the Pseudomonas quinolone signal (PQS, 2-heptyl-3-hydroxy-4(1H)-quinolone), which plays a central role in the regulation of virulence factor production by Pseudomonas aeruginosa. We present here the crystal structures of AqdC in its native state and in complex with the PQS cleavage product N-octanoylanthranilic acid, and of mutant AqdC proteins in complex with PQS. AqdC possesses an alpha/beta-hydrolase fold core domain with additional helices forming a cap domain. The protein is traversed by a bipartite tunnel, with a funnel-like entry section leading to an elliptical substrate cavity where PQS positioning is mediated by a combination of hydrophobic interactions and hydrogen bonds, with the substrate's C4 carbonyl and C3 hydroxyl groups tethered by His97 and the catalytic His246, respectively. The side chain of the AqdC-bound product extends deeper into the "alkyl tail section" of the tunnel than PQS, tentatively suggesting product exit via this part of the tunnel. AqdC prefers PQS over congeners with shorter alkyl substituents at C2. Kinetic data confirmed the strict requirement of the active-site base His246 for catalysis, and suggested that evolution of the canonical nucleophile/His/Asp catalytic triad of the hydrolases to an Ala/His/Asp triad is favorable for catalyzing dioxygenolytic PQS ring cleavage.
ESTHER : Wullich_2019_J.Struct.Biol_207_287
PubMedSearch : Wullich_2019_J.Struct.Biol_207_287
PubMedID: 31228546
Gene_locus related to this paper: mycab-x8en65