Objetivos

The lipid structure is characterised by the presence of fatty acid moieties. The

physical and chemical characteristics of individual fats and oils are determined mainly by their fatty acid composition. The fatty acids in biological systems are all straight- chain compounds, usually containing an even number of carbon atoms, typically ranging from fourteen to twenty four. Their main structural characteristics are chain

length with a presence of double bonds (unsaturation) and of substituent groups. They are rarely found free in nature but occur in esterified forms as the major components of the various lipids which mostly with glycerol. The predominant fatty acids in

higher plants and animals are these with sixteen or eighteen carbon units: palmitic (16:0), oleic (18:1), linoleic (18:2) and stearic (18:0) acid. Fatty acid with less than

fourteen or more than twenty carbon atom are less common. In addition, most fatty acids have an even number of carbon atoms because they are usually biosynthesised

by the condensation of two carbon units. More than half of the fatty acid residues of plant and animal lipids are unsaturated, and often polyunsaturated. The C^g

polyunsaturated fatty acids, such as oleic, linoleic and linolenic acids, are important as

major components of most plant lipids including commercially important vegetable oils. They are essential because they cannot be synthesised by animal tissues, and are

biosynthetic precursors in the synthesis of larger fatty acid polymers (Stryer, 1995;

Vance and Vance, 1991).

INTRODUCTION Lipids

1.4.1.1

Catabolism of fatty acids

The ability to utilise fatty acids, oils and fats is found widely amongst bacteria, yeasts and moulds. The ester link between the glycerol backbone and fatty acid is

hydrolysed by an intracellular or extracellular lipase enzyme (Section 1.3). The

glycerol is then brought to the Embden-Myerhoff pathway and is converted to a

glycolytic intermediate later in the oxidation cycle, while fatty acids, which are

extremely toxic to the cell, are processed through oxidation cycle. However, fatty

acids are not transferred directly to the oxidation cycle, but must be transformed into the thiol esters linked with the complex nucleotide coenzyme A (Light, 1969). The

passage through the cytoplasmic membrane is less clear. Possibly, acyl-CoA synthetase is loosely associated with the inner side of the cytoplasmic membrane and immediately activates entering fatty acids to avoid the unwanted detergent effect

(Fuchs, 1999). Generally, the process is catalysed by a family of at least three acyl- CoA synthetase that differ according to their chain length specificity. In addition, the pyrophosphatase reaction (Reaction 1.3) renders the reversible reaction (Reaction 1.2) irreversible; two high-energy phosphate equivalents are required. Medium-chain (C7-

Cji) fatty acids, which are more toxic, may diffuse and then become activated (Fuchs,

1999). These steps are shown in the following reactions (Boyer, 1983):

R-fCHJa-COOH + CoA-SH + ATP Acyl-CoA-synthetase ^ R_(CH2)3-CO-SCoA + AMP + PP;

Fatty Acyl CoA (1 .2 )

PP, + HjO 2 P, (1.3)

There are four reactions (Reaction 1.4-1.7) take place during the degradation of

CoA-activated fatty acids. The mechanism used for breaking down the molecule of fatty acyl CoA known as P-oxidation because the carbon P to the activated carbon is

prepared to be oxidised. The mechanisms are as follows (Fuchs, 1999; Stryer, 1995):

(1) Formation of a rm»j-double bond by acyl-CoA-dehydrogenase.

R-(CH7)3-CO-SCoA+FAD ^ rch^CH=CH-CO-SCoA+FADH; (1 .4 )

Fatty Acyl CoA Enoyl-CoA

INTRODUCTION Lipids

(2) Hydration of the double bond by enoyl-CoA-hydratase.

RCH2CH=CH-CO-SCoA + H2O Enoyl-CoA-hydrat^e _ ^ RCH2CH(OH)-CH2CO-SCoA ( 1 .5 )

Enoyl CoA L-hydroxyacyl CoA

(3) Dehydrogenation of the p-hydroxyl group (formed in the previous step) by 3-L- hydroxyl-acyl-CoA-dehydrogenase to form P-ketoacyl-CoA.

RCH2CH(OH)-CH2CO-SCoA + NAD^ + ^-L-hydroxyacyl-CoA-dehydrogenase ^ (1 .6 )

L-hydroxyacyl CoA RCH2CO-CH2CO-SC0A + NADH +

fi-ketoacyl CoA

(4) a -C and P~C cleavage in a thiolysis reaction with coenzyme A catalysed by P~ ketoacyl-CoA-thiolase (thiolase) to form acetyl-CoA and a new acyl-CoA containing two less carbon atom than the fatty acyl CoA present in the first step of the cycle.

RCH2CO-CH2CO-SC0A + CoASH P-ketoacyl-CoA-thiolase ^ cHjCO-SCoA + RCH2-CO-SC0A *

P-ketoacyl CoA Acetyl CoA Fatty Acyl CoA

( 1 .7 )

* This intermediate obtains 2 carbons less than the substrate in step (1), and will go back to serve as a substrate in step (1) and continue the subsequent rounds until 2

molecules of acetyl CoA produced at the last round. Acetyl CoA generated from each round will serves as an initial substrate in TCA cycle (Stryer, 1995a).

For example, the degradation of palmitoyl CoA (Cjg-acyl CoA) requires seven

reaction cycles. In the seventh cycle, the C^-ketoacyl CoA is thiolysed to two molecules of acetyl CoA. Hence, the stoichiometry of oxidation of palmitoyl CoA is

Palmitoyl CoA + 7 FAD + 7 NAD+ + 7 CoA + 7 H2O --- > ( 1 .8 )

8 acetyl CoA + 7 FADH2 + 7 NADH + 7 H^

The net reaction of the citric acid cycle is (Stryer, 1995a)

A cetyl CoA + 3 NAD+ + FAD + GDP + P, + 2 H2O --- > ( 1 .9 )

2 CO2 + 3 NADH + FADH2 + GTP + 2H+ + CoA

Most fatty acids have even number of carbon atoms which can be utilised completely

to acetyl CoA while odd number fatty acid which normally found in some plants and microorganisms gives propionyl CoA at the last cycle which is converted to succinyl

CoA, via the form of methylmalonyl CoA for entry into the citric acid cycle.

Furthermore, unsaturated fatty acid from biological origin contains only cis- double bonds which often begin between C9 and CIO.

INTRODUCTION Lipids

After the oxidation cycle the double bond ends in being between C3 and C4 which

enzyme isomerase then converts the cis-2> into a trans-2 which the enoyl-CoA- hydratase, o f the second reaction, can continue the process on the pathway (Numa,

1984).

1.4.2 Application of oils in fermentation (antibiotic production)