Nonenzymic ADP-ribosylation

Hilz H, Koch R, Fanick W, Klapproth K, Adamietz P (1984) Nonenzymic ADP-ribosylation of specific mitochondrial polypeptides. Proc Natl Acad Sci USA 81 3929-3933... 

Nonenzymic ADP-Ribosylation and Phospho(ADP-Ribosylation) of SpecMc Acceptor Proteins... 

Mitochondrial ADP-ribosylation as described by Kun et al. [2,4] involves the apparent transfer of ADPR residues from NAD to a 50 kD polypeptide. Yet another ADP-ribosylation reaction was reported by Richter et al. [13,14], They described the modification of a 30 kD polypeptide insubmitochondrial particles when incubated with labeled NAD. This reaction was brought into connection with Ca efflux from mitochondria as induced by treatment of mitochondria with organic peroxides. Such treatment was known to result in the concomitant degradation of NAD and NADP to ADPR and P-ADPR, respectively [15]. During our attempts to characterize these mitochondrial systems, evidence accumulated that nonenzymic ADP-ribosylation is involved. The following observations support this interpretation ... 

In alkylated cells considerable amounts of free ADPR must be formed in consequence of the highly stimulated turnover of (ADPR)j, proteins [3,4]. Since free ADPR can form acid-insoluble protein conjugates [7, 8], mono(ADP-ribosyl) histone HI could have been arisen by such a reaction. We, therefore, analyzed endogenous conjugates with respect to chemical stability and compared it with other conjugates of known structure as well as with nonenzymic ADPR-Hl adducts. In addition, location of the ADPR groups on the histone HI molecule was investigated. 

Extension of these studies to the stability pattern of other enzymic and nonenzymic ADPR conjugates (unpubhshed experiments) revealed that ADP-ribosyl... 

To test these possibilities, the histone HI fraction was purified from alkylated cells and subjected to the stability treatments described above. Chemical stability of the endogenous material was different in all aspects from ADPR-histone HI formed in vitro by nonenzymic reaction and from gju- or arg-linked ADPR conjugates. From our data, we conclude that in alkylated hepatoma cells, mono(ADP-ribosyl) groups in HI are neither linked by an ester group, nor via arginine residues. They also do not represent products of nonenzymic reaction. 

The alkylation-induced changes had demonstrated that important acceptors like histone HI may become nearly exclusively mono(ADP-ribosyl)ated. Mono(ADP-ribosyl)ation of proteins also appears to be of importance in certain diffemtiation processes. When F9 teratocarcinoma cells were incubated with low concentrations of retinoic acid, they differentiated into endoderm cells [22]. Under these conditions, mono(ADPR) conjugates increased considerably. The increase was time dependent, and it was mainly due to a fraction that was neither carboxyl- nor arginine-linked, nor did it correspond to nonenzymic conjugates. However, further experiments are required to determine the nature of these conjugates. 

Neither transfer of single ADPR residues by specific eukaryotic transferases nor the existence of other than ester-type linkages in ADPR protein conjugates were recognized for a number of years [24]. Now, there is increasing evidence that the eukaryotic cell possesses a rather elaborate instrumentarium of ADP-ribosylation reactions to modulate important processes. The use of mono(ADPR) groups for the covalent modification of protein provides a tool that may prove to be as versatile as poly(ADP-ribosyl)ation. In addition to the systems discussed here (alkylated cells, differentiation, tumor promotors), mono(ADP-ribosyl)ation may be involved also in the modulation of protein synthesis [8], in the response to heat shock (M. Jacobson, personal communication), and in glutamine synthesis [25]. Nonenzymic mono(ADP-ribosyl)ation reactions must also be taken into consideration whenever the functions of active (P)ADP-ribose as present in the pyridine nucleotides are studied. Finally, preliminary data may indicate that an additional way to modulate protein functions could be provided by phospho-(ADP-ribosylation) reactions. 

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