Modification by enzymes

An alternative to modifying the functional group attached to fibrils is to utilise the chemistry present in the amino acid side chains. Furthermore, as peptides often undergo specific modification by enzymes in vivo, these could be harnessed for synthetic purposes. Qll (Ac-QQKFQFQFEQQ-Am, a fibril-forming peptide based on Pi 1-2), was coupled to lysine-based molecules by treatment with an enzyme (tissue transglutaminase, TGase) which results in a reaction between lysine and glutamine side chains [72] (Fig. 32). 

The serine residue of isocitrate dehydrogenase that is phos-phorylated by protein kinase lies within the active site of the enzyme. This situation contrasts with most other examples of covalent modification by protein phosphorylation, where the phosphorylation occurs at a site remote from the active site. What direct effect do you think such active-site phosphorylation might have on the catalytic activity of isocitrate dehydrogenase (See Barford, D., 1991. Molecular mechanisms for the control of enzymic activity by protein phosphorylation. Bioehimiea et Biophysiea Acta 1133 55-62.)... 

Phosphonate analogs to phosphate esters, in which the P—0 bond is formally replaced by a P—C bond, have attracted attention due to their stability toward the hydrolytic action of phosphatases, which renders them potential inhibitors or regulators of metabolic processes. Two alternative pathways, in fact, may achieve introduction of the phosphonate moiety by enzyme catalysis. The first employs the bioisosteric methylene phosphonate analog (39), which yields products related to sugar 1-phosphates such as (71)/(72) (Figure 10.28) [102,107]. This strategy is rather effective because of the inherent stability of (39) as a replacement for (25), but depends on the individual tolerance of the aldolase for structural modification close... 

Martirosova, E., Karpekina, T., El -Registan, G. Enzyme modification by natural chemical chaperons of microorganisms. Microbiology, Vol.73, No.5, (August 2004), pp. 609-615, ISSN 1350-0872... 

It is not only the activity that can be altered by incorporation of noncoded amino acids. Introduction of structures possessing certain chemical functions leads to the possibility of highly regioselective modification of enzymes. For example, selective enzymatic modification of cystein residues with compounds containing azide groups has led to the preparation of enzymes that could be selectively immobilized using click chemistry methods [99]. 

In mammalian cells, the two most common forms of covalent modification are partial proteolysis and ph osphorylation. Because cells lack the ability to reunite the two portions of a protein produced by hydrolysis of a peptide bond, proteolysis constitutes an irreversible modification. By contrast, phosphorylation is a reversible modification process. The phosphorylation of proteins on seryl, threonyl, or tyrosyl residues, catalyzed by protein kinases, is thermodynamically spontaneous. Equally spontaneous is the hydrolytic removal of these phosphoryl groups by enzymes called protein phosphatases. 

Ghanges in the availability of substrates are responsible for most changes in metabolism either directly or indirectly acting via changes in hormone secretion. Three mechanisms are responsible for regulating the activity of enzymes in carbohydrate metabolism (1) changes in the rate of enzyme synthesis, (2) covalent modification by reversible phosphorylation, and (3) allosteric effects. 

Chromatin remodeling, transcription factor modification by various enzyme activities, and the communication between the nuclear receptors and the basal transcription apparatus are accomplished by protein-protein interactions with one or more of a class of coregulator molecules. The number of these coregulator molecules now exceeds 100, not counting species variations and splice variants. The first of these to be described was the CREB-binding protein, CBP. CBP, through an amino terminal domain, binds to phosphorylated serine 137 of CREB and mediates transactivation in response to cAMP. It thus is described as a coactivator. CBP and... 

Mechanisms of lipid peroxidation that have been implicated in atherosclerosis may be pertinent to RA. Cellular lipoxygenase enzymes may promote LDL modification by inserting hydroperoxide groups into unsaturated fetty-acid side chains of the LDL complex (Yla-Herttuala etal., 1990). 15-Lipoxygenase has been implicated as an initiator of LDL oxidation (Cathcart etal., 1991) whilst 5-lipoxygenase does not appear to be involved (Jessup et al., 1991). Products of activated lipoxygenase enzymes within inflammatory synovial fluid surest that this pathway could be activated in RA (Costello etal., 1992). 

Note that in some cases one may follow the time course of covalent E-A formation by equilibrium binding methods (e.g., LC/MS, HPLC, NMR, radioligand incorporation, or spectroscopic methods) rather than by activity measurements. In these cases substrate should also be able to protect the enzyme from inactivation according to Equation (8.7). Likewise a reversible competitive inhibitor should protect the enzyme from covalent modification by a mechanism-based inactivator. In this case the terms. S and Ku in Equation (8.7) would be replaced by [7r] and K respectively, where these terms refer to the concentration and dissociation constant for the reversible inhibitor. 

Virus restriction and modification by the host We have already seen that one form of host resistance to virus arises when there is no receptor site on the cell surface to which the virus can attach. Another and more specific kind of host resistance involves destruction of the viral nucleic acid after it has been injected. This destruction is brought about by host enzymes that cleave the viral DNA at one or several places, thus preventing its replication. This phenomenon is called restriction, and is part of a general host mechanism to prevent the invasion of foreign nucleic acid. 

UV-C technology is widely used as an alternative to chemical sterilization and microorganism reduction in food products (Lamikanra 2002 Fan and others 2008). Ultraviolet light also induces biological stress in plants and defense mechanisms in plant tissues with the consequent production of phytochemical compounds (Lee and Kader 2000). Phytoalexin accumulation could be accompanied by other inducible defenses such as cell-wall modifications, defense enzymes, and antioxidant activity, which have been reported with health benefits (Gonzalez-Aguilar and others 2007). It is well documented that UV-C irradiation has an effect in secondary metabolism. 

For enzymic assays, a modification by Voragen and coworkers is recommended.215... 

The catalytic activity of cPLA2 is stimulated by phosphorylation catalyzed by the mitogen-activated protein kinase (MAPK) at Ser505. This modification stimulates enzyme activity only, indicating that translocation and phosphorylation are independent mechanisms of cPLA2 regulation [21]. 

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