Trypanosoman makrofomeeni

Trypanosoma crusei
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4827093/

Sci Rep. 2016; 6: 24213.

Published online 2016 Apr 11. doi: 10.1038/srep24213

PMCID: PMC4827093

PMID: 27064071

Proximal ADP-ribose Hydrolysis in Trypanosomatids is Catalyzed by a Macrodomain

Teemu Haikarainen¹ and Lari Lehtiö^a,1

Abstract

ADP-ribosylation is a ubiquitous protein modification utilized by both prokaryotes and eukaryotes for several cellular functions, such as DNA repair, proliferation and cell signaling. Higher eukaryotes, such as humans, utilize various enzymes to reverse the modification and to regulate ADP-ribose dependent signaling. In contrast, some lower eukaryotes, including trypanosomatids, lack many of these enzymes and therefore have a much more simplified ADP-ribose metabolism. Here we identified and characterized ADP-ribose hydrolases from Trypanosoma brucei and Trypanosoma cruzi, which are homologous to human O-acetyl-ADP-ribose deacetylases MacroD1 and MacroD2. The enzymes are capable for hydrolysis of protein linked ADP-ribose and a product of sirtuin-mediated lysine deacetylation, O-acetyl-ADP-ribose. Crystal structures of the trypanosomatid macrodomains revealed a conserved catalytic site with distinct differences to human MacroD1 and MacroD2.

 ADP-ribosylation 
is a covalent modification where one (mono) or multiple  (poly) ADP-ribose units are attached to a target protein. In eukaryotes, the modification is catalyzed by poly-ADP-ribose polymerases (PARPs), silent information regulators (sirtuins) and poorly characterized membrane anchored arginine ADP-ribosyltransferases (ARTs)^1,2,3.

 ADP-ribosylation regulates several cellular events, including DNA repair, cell cycle progression, transcription and cell death. In humans over 20 enzymes have been found to catalyze ADP-ribosylation and modification can be reversed by various enzymes:
 PARG (poly(ADP-ribose) glycohydrolase) and
 ARH3 (ADP-ribosylhydrolase 3), which can hydrolyze ADP-ribose polymer leaving the proximal ADP-ribose molecule 8mono-ADP-ribosyl)attached to the modified protein;
MacroD1 and MacroD2, which cleave the proximal mono-ADP-ribosyl;
 and OARD1 (O-acetyl-ADP-ribose deacetylase 1), which is capable of hydroxylation of the proximal mono-ADP-ribosylation and also cleaving the entire PAR en bloc⁴.
There are also less characterized enzymes involved in ADP-ribose hydrolysis, namely ARH1, which hydrolyzes mono-ADP-ribosylated arginines⁵ and Nudix hydrolase 16, which is able to remove both mono-and poly-ADP-ribosylation leaving the modified protein with a ribose-5′-phosphate⁶. MacroD1, MacroD2, OARD1 and ARH3 have also O-acetyl-ADP-ribose hydrolysis activity as they are capable of removing the acetyl group from O-acetyl-ADP-ribose produced by sirtuin-mediated lysine deacetylation^7,8,9.


( Merkitystä trypanosomassa:)
 Trypanosoma brucei and Trypanosoma cruzi are parasitic protozoa responsible for severe human and animal diseases. T. brucei is the causative agent of African trypanosomiasis or sleeping sickness and T. cruzi is responsible for South American trypanosomiasis or Chagas Disease.
 These parasites seem to have a highly simplified ADP-ribose metabolism compared to higher eukaryotes, such as humans.
They contain a single PARP opposed to 17 PARPs found in humans^10,11,12 and have two (T. cruzi) or three (T. brucei) sirtuins, while humans have seven (although not all have ADP-ribosylating activity)¹³.
Only one ADP-ribose hydrolase, PARG, has been characterized from the parasites¹⁴.

 Recently, a phylogenetic analysis of proteins linked to ADP-ribose metabolism identified a single MacroD1/MacroD2 homologue in T. brucei¹⁵. We analyzed the available genome sequences and observed that besides PARG and MacroD1/MacroD2 homologue, T. brucei and T. cruzi do not have other known enzymes capable for the hydrolysis of ADP-ribose. We show that the trypanosomatid MacroD1/MacroD2 homologues are hydrolyzing both protein-linked ADP-ribose and free O-acetyl-ADP-ribose (O-AADPR) The crystal structures of the enzymes reveal highly conserved macrodomain folds containing conserved ADP-ribose binding sites.

Results

Identification and characterization of trypanosomal proximal ADP-ribose hydrolases

ADP-ribose hydrolyzing enzymes of T. cruzi and T. brucei were searched from NCBI non-redundant database using known ADP-ribose hydrolases from human (PARG, ARH3, ARH1, OARD1, MacroD1, MacroD2 and Nudix hydrolase 16) as a query (Fig. 1A).
 Only one putative ADP-ribose hydrolase was identified from the parasites, which had homology to MacroD1 and MacroD2 (Fig. 1B). We named these proteins as Trypanosoma brucei MacroD-like protein (TbMDO) and Trypanosoma cruzi MacroD-like protein (TcMDO). It should be noted that T. brucei and T. cruzi contain putative Nudix hydrolases but they have very low identities to human Nudix hydrolase 16 and are more similar to other human Nudix proteins.
Human MacroD1 and MacroD2 function as ADP-ribose hydrolases cleaving ADP-ribose from target proteins, as well as O-acetyl-ADP-ribose deacetylases hydrolyzing O-acetyl-ADP-ribose produced by sirtuins during lysine deacetylation (Fig. 2A).