Butyryl unit

Fatty acids are synthesized by a multistep route that starts with acetate. The first step is a reaction between protein-bound acetyl and malonyl units to give a 3-keto-butyryl unit. Show the mechanism, and tell what kind of reaction is occurring. (See Section 29.7). 

Fig. 6. Conversion of weight percent to degree of substitution per anhydroglucose unit (max = 3.0), where is butyryl, is propionyl, and 0 is acetyl.

The net result of this biosynthetic cycle is the synthesis of a four-carbon unit, a butyryl group, from two smaller building blocks. In the next cycle of the process, this butyryl-ACP condenses with another malonyl-ACP to make a... 

The net effect of the eight steps in the fattv-acid biosynthesis pathway is to take two 2-carbon acetyl groups and combine them into a 4-carbon butyryl group. Further condensation of the butyryl group with another malonyl AGP yields a 6-carbon unit, and still further repetitions of the pathway add two more carbon atoms to the chain each time until the 16-carbon palmitoyl ACP is reached. 

Tacrine is a non-competitive, irreversible inhibitor of both acetyl and butyryl cholinesterase, with a greater potency for the latter enzyme. Based on the outcome of placebo-controlled, double-blind studies, tacrine was the first anticholinesterase to be licensed for the symptomatic treatment of AD in the United States. The main disadvantage of tacrine lies in its hepatotoxicity (approximately 50% of patients were found to develop elevated liver transaminases which reversed on discontinuation of the drug). Because of such side effects and limited efficacy, tacrine is no longer widely prescribed. 

In applying mutasynthesis to the production of novel avermectins, the elimination of branched-chain a-keto acid dehydrogenase (BCDH) was targeted. This multienzyme complex is responsible for supplying the 2-methylbutyryl- and iso-butyryl-CoA starter units that initiate natural avermectin biosynthesis [21,29],... 

The DEBS 1-TE multienzyme was purified to 90-95% homogeneity and then used in another series of experiments to establish the extent to which alternative starter units could be used by the polyketide synthase [36], Substantial amounts of lactones were obtained in the presence of acetyl-, n-butyryl-, and isobutyryl-CoA, illustrating that the loading didomain exhibits a relaxed specificity for the starter unit (Fig. 10). The utilization of acetyl-CoA and -butyryl-CoA by DEBS 1 + TE was demonstrated in a cell-free system [39], Additionally, in the absence of the reducing cofactor NADPH, cell-free DEBS 1+TE converted... 

Figure 10 Biosynthesis of triketide lactones in vitro by DEBS 1-TE. Incubation of propionyl-CoA or acetyl-CoA with methylmalonyl-CoA and NADPH in vitro resulted in synthesis of the appropriate lactones. The mini-PKS also accepted the unnatural starter units ra-butyryl-CoA and isobutyryl-CoA.

Figure 17 Design of assays to evaluate decarboxylation of methylmalonyl-CoA catalyzed by DEBS 1-TE. It was anticipated that a decarboxylase activity would incorporate deuterium into the starter unit of propionyl lactone and that such labeling would be visible by GC-MS analysis. As decarboxylation has been reported in the presence of primers other than propionyl-CoA, assays I—III included n-hutyryl-CoA as a starter unit. Assay IV was designed to evaluate the possibility that -butyryl-CoA suppresses decarboxylation. GC-MS analysis gave no evidence for labeling of the side chain in any assay. Therefore, decarboxylation is not a significant reaction of KSt under these conditions.

The pathway The first committed step in fatty acid biosynthesis is the carboxylation of acetyl CoA to form malonyl CoA which is catalyzed by the biotin-containing enzyme acetyl CoA carboxylase. Acetyl CoA and malonyl CoA are then converted into their ACP derivatives. The elongation cycle in fatty acid synthesis involves four reactions condensation of acetyl-ACP and malonyl-ACP to form acetoacetyl-ACP releasing free ACP and C02, then reduction by NADPH to form D-3-hydroxybutyryl-ACP, followed by dehydration to crotonyl-ACP, and finally reduction by NADPH to form butyryl-ACP. Further rounds of elongation add more two-carbon units from malonyl-ACP on to the growing hydrocarbon chain, until the C16 palmitate is formed. Further elongation of fatty acids takes place on the cytosolic surface of the smooth endoplasmic reticulum (SER). 

This first round of elongation produces the four-carbon butyryl-ACP. The cycle now repeats with malonyl-ACP adding two-carbon units in each cycle to the lengthening acyl-ACP chain. This continues until the 16-carbon palmitoyl-ACP is formed. This molecule is not accepted by the acyl-malonyl-ACP condensing enzyme, and so cannot be elongated further by this process. Instead it is hydrolyzed by a thioesterase to give palmitate and ACP. 

DEBS appears to have a fairly broad starter unit specificity. Butyryl-CoA and acetyl-CoA are incorporated into the respective Ciq- and Cg-lactones, although at significantly lower yields than propionyl-CoA [160,161] (Fig. 9). Remarkably, DEBS 1 -I- TE can process unreduced and partially reduced intermediates as well. The AT-L in the NH2-terminal part of DEBS 1 covalently activates the starter units with comparable efficiency [161]. In contrast, malonyl CoA is not recognized as an extender unit by acyltransferase domains (R. Pieper, unpublished). When NADPH is excluded from the reaction mixture, a pyran-2-one was synthesized by DEBS 1 -I- TE [ 161 ]. Consistent with this result, incubation of (2S, 3R)-2-methyl-hydroxypentanoyl-NAC thioester and methylmalonyl CoA with DEBS... 

The overall for the synthesis of the Cg-lactone by DEBS 1+TE was measured to be 3.4 min [163]. For 6-dEB synthesis by complete DEBS, an apparent /Cjat of 0.5 min was determined. The measured of DEBS 1+TE indicates that this truncated PKS is highly active in a cell-free system and approaches catalytic activity comparable to in vivo levels. The apparent for (2S) methylmalonyl CoA consumption by DEBS 1+TE is 24 pM. Although starter units with shorter and longer side chains are incorporated into the respective triketide 6 lactones, DEBS 1+TE has a 7.5-fold preference for propionyl-CoA over butyryl-CoA and a 32-fold preference over acetyl-CoA. In the absence of the primer propionyl-CoA, DEBS 1+TE turns over (2S)-methylmalonyl CoA and NADPH at the same rate as in the presence of the starter unit. This suggests that DEBS 1+TE decar-boxylates the extender unit, transfers the evolving propionyl group to its active site KSl, and thus initiates polyketide chain elongation. 

Fatty acids have predominantly even numbers of carbon atoms because they are effectively formed from acetyl (C2) units, which are derived from glucose in the presence of various enzymes, coenzymes and carrier proteins. An overall scheme for saturated fatty acid biosynthesis is presented in Fig. 2.13, in which it can be seen that the first step is the formation of acetyl coenzyme A (often abbreviated to acetyl-CoA). One molecule ofacetyl-CoA undergoes addition of CO, to form malonyl-CoA, while the acetyl group on another molecule is transferred to an enzyme (fatty acid synthase). The malonyl unit (C3) is added to the enzyme-bound acetyl unit, which produces a butyryl group following loss of C02, dehydration and reduction. Six further steps of combined malonyl addition, decarboxylation, dehydration and reduction occur to yield palmitate (C16). Higher acids are built from palmitate in a similar... 

Fig. 2.13 Biosynthesis of saturated fatty acids in plants and animals. Palmitate is formed by successive additions of malonyl coenzyme A to the enzyme-bound chain, with C02 being lost at each addition.This results in chain elongation by a (CH2)2 unit at each step. Details of the formation of butyryl (C4) from acetyl (C2) are shown, while the subsequent six further additions, terminating in palmitate, proceed similarly.

ACP, with subsequent dehydration to enoyl-ACP, which is further reduced to butyryl-ACP (Voelker Kinney, 2001). Later steps of 2-carbon unit additions are catalyzed by KAS I (Shimakata Stumpf, 1983). Once palmitoyl-ACP is formed, KAS II may add an extra 2-carbon unit to form stearoyl-ACP (Shimakata Stumpf, 1982). The formation of oleoyl-ACP is catalyzed by stearoyl-ACP desaturase, an enzyme that promotes the formation of double bonds in the position (Shanklin Gaboon, 1998). This enzyme also desaturates palmitoyl-ACP in position A . However, the apparent specificity factor for stearoyl-ACP is much greater than for palmitoyl-ACP (Gibson, 1993). The presence of two A stearoyl-ACP desaturases coded by two different genes has been reported in soybean (Byfield et al., 2006). 

Polyketides. Collective name for natural products produced biosynthetically by way of poly(/5-oxo-carboxylic acids). The name was derived in 1907 by Collie on the basis of the hypothesis that natural prt ucts may be formed by multiplication of ketene (HjCsC o) units. The P. chains are constructed on multienzyme complexes (polyketide synthases) from acetyl- and malonyl-CoA ( acetogenins) or also by use of propio-nyl- and/or butyryl-CoA. Depending on the number of building blocks the natural products ate classified as triketides (n=3), tetraketides (n=4), etc. 

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