Transferase reactions

Perhaps the most significant case of catalysis by RNA occurs in protein synthesis. Harry F. NoIIer and his colleagues have found that the peptidyl transferase reaction, which is the reaction of peptide bond formation during protein synthesis (Figure 14.24), can be catalyzed by 50S ribosomal subunits (see Chapter 12) from which virtually ail of the protein has been removed. These... 

Iordanov, M. S. et al. Ribotoxic stress response Activation of the stress-activated protein kinase JNK1 by inhibitors of the peptidyl transferase reaction and by sequence-specific RNA damage to the alpha-sarcin/ricin loop in the 28S rRNA. Mol. Cell. Biol. 17, 3373, 1997. 

The origin of the idea that a ribosome might be a ribozyme is derived from the experiment in which peptidyl transferase activity was observed even after digestion of protein components of the ribosome [15]. This was surprising because the most important biological function involved in the synthesis of proteins is catalyzed by RNA. Recently, a large ribosomal subunit from Haloarcula marismortui was determined at a resolution of 2.4 A [16, 155]. Importantly, because of the absence of proteins at the active site, it was concluded that the key peptidyl transferase reaction is accomplished by the ribosomal RNA (rRNA) itself, not by proteins. How does it work ... 

Histone acetyltransferases (H ATs) catalyze the transfer of an acetyl moiety from acetyl-CoA to the E-amino group of certain lysine residues within core histone proteins. This transferase reaction produces acetylated histones and the deacetylated cofactor CoA-SH. As HATs are important enzymes in the regulation of gene expression, there are also a number of assays available to detect acetyltransferases activity. 

Large amounts of pentachlorothiophenol were formed during the coupled reaction. Apparently the methyl transferase reaction was rate-limiting in the coupled system. 

The methyl transferase reaction was demonstrated with penta-chlorothiophenol and a number of other substrates. The importance of this reaction in pesticide metabolism needs further evaluation. Some conversion of pentachlorothioanisole to pentachlorothioanisole sulfoxide was observed vivo, but this reaction was not specifically Investigated. 

Evidence was also provided that Insoluble residues may be produced from GSH conjugates via cysteine conjugate or thiol Intermediates. These studies also suggested that certain reactions should be studied in greater detail to assess their importance in pesticide metabolism l.e., the C-S lyase reaction, the methyl transferase reaction, and the transamination reaction. 

Chloramphenicol is able to inhibit the peptidyl transferase reaction and so bacterial protein synthesis by binding reversibly to the 50s ribosomal subunit. Resistance can occur due to the plasmid-mediated enzyme chloramphenicol acetyltransferase which inactivates the drug by acetylation. Such resistance is often a part of plasmid-mediated multidrug resistance. Resistance can also occur by an altered bacterial permeability. However in most instances resistance to chloramphenicol only develops slowly and remains partial. 

Lipases can be used in transferase reactions to exchange fatly acids in fats. This is of considerable interest to the food industry. The enzymatic production of cocoa butter substitutes is the most well-known example. Cocoa butter is the fat component in chocolate. It melts in the range between room temperature and body temperature because its triglyceride molecules contain certain combinatiorts of fatly acids. Natiual... 

Another type of important selectivity is that between hydrolysis and transferase reactions (transesterification, transglycosylation, etc.) catalyzed by hydrolases. In this case, water can act both as a reactant and as a substance that modifies the properties of the enzyme. Effects of water as a reactant can be expected to be governed by the concentration or activity of water, as with other substrates. The effects of water as an enzyme modifier are considerably more difficult to predict. 

The most straightforward way to quantify the competition between the transferase reaction and hydrolysis is to measure the initial ratio of these two reactions. Intuitively, one would assume the transferase/hydrolysis ratio to decrease with increasing water activity because of the effect of water as a reactant This is often the case when lipases are used as catalysts [34—36]. However, in reactions catalyzed by glycosidases and proteases the transferase/hydrolysis ratio can either increase or decrease with increasing water activity [37, 38]. 

Smart, J.B. (1993) Transferase reactions of /3-galactosidases - New product opportunities, in Lactose Hydrolysis, Bulletin 239, International Dairy Federation, Brussels, pp. 16-22. 

S-adenosylmethionine is also a biological methyl group donor. The product of its methyl transferase reactions is S-adenosylhomocysteine. This product is further degraded by S-adenosylhomocysteine hydrolase, an enzyme that contains tightly bound NAD+, to form homocysteine and adenosine. 

Since preliminary studies showed that 6-hydroxymellein-O-methyl-transferase activity was appreciably inhibited in the presence of the reaction products, the mode of product inhibition of the enzyme was studied in detail in order to understand the regulatory mechanism of in vivo methyltransfer. It is well known that S-adenosyl-Z.-homocysteine (SAH), which is a common product of many O-methyltransferases that use SAM as methyl donor, is usually a potent inhibitor of such enzymes. In the 6-hydroxymellein-Omethyltransferase catalyzing reaction another product of this enzyme, 6-methoxymellein, has pronounced inhibitory activity, in addition to SAH. Since the specific product of the transferase reaction, 6-methoxymellein, is capable of inhibiting transferase activity [88], this observation suggests that activity of the transferase is specifically regulated in response to increases in cellular concentrations of its reaction products in carrot cells. It has been also found that 6-methoxymellein inhibits transferase activity with respect not only to 6-hydroxymellein but also to SAM, competitively. This competitive inhibition was also found in SAH as a function of the co-substrates of the enzyme [89]. It follows that the reaction catalyzed by 6-hydroxymellein-O-methyltransferase proceeds by a sequential bireactant mechanism in which the entry of the co-substrates to form the enzyme-substrate complexes and the release of the co-products to generate free enzyme take place in random order [Fig. (7)]. This result also implies that 6-methoxymellein and SAH have to associate with the free transferase protein to exhibit their inhibitory activities, and cannot work as the inhibitors after the enzyme forms complexes with the the substrate. If, therefore, 6-hydroxymellein-O-methyltransferase activity is controlled in vivo by its specific product 6-methoxymellein, this compound should... 

Thus tris is about 100 times better as an acceptor than water. It was later shown that as with other phosphatases (140) many compounds can participate as active acceptors in transferase reactions (see Table IX) (123, 124). Other compounds, not shown in Table IX, which show transferase activity are L-glucose, glucosamine, and butanolamine (124). Compounds which do not show exceptionally marked transferase ability are sodium lactate, fluoroethanol, ethanol, ethylenediamine, catechol, 2-... 

Slight activation by pyruvate phos-phoenolpyruvate a competitive inhibitor of phosphohydrolase activity and also a phosphoryl donor in the transferase reaction... 

Biosynthesis of farnesyl pyrophosphate. Farnesyl pyrophosphate is a C 5 intermediate containing three C5 isoprenoid subunits. The two transferase reactions involved in the formation of farnesyl pyrophosphate occur by virtually identical mechanisms as shown. 

One inhibitor of those shown in table 23.3 specifically interferes with a step in pyrimidine nucleotide biosynthesis. Af-(Phosphonacet T)-L-aspartate (PALA) is a powerful inhibitor of the carbamoyl transferase reaction. PALA was synthesized to act as an analog of the transition state intermediate (see chapter 9) postulated to be formed in the aspar-... 

Figure 7.19 Glutathione transferase reaction and formation of mercapturic acids.

It is probable that methyl transferase reactions proceed by nucleophilic attack on S-adenosylmethionine94 and may involve an SN2-like transition state.95 Thus, inversion of configuration as observed in indolmycin biosynthesis indicates that an odd number of nucleophilic displacements occurs and suggests that the methyl group is transferred directly from donor to (112), i.e. without generation of a methylated-enzyme intermediate.92 The combined results are summarized in Scheme 11. 

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