Humboldt State University ® Department of Chemistry

Richard A. Paselk

Chem 438 - Introductory Biochemistry - Spring 2013

Lecture Notes:

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Fatty Acid Biosynthesis, cont.

Fatty Acid Desaturation, cont.

Both plants and animals use mixed function oxidases (simultaneously oxidize two substrates): Acyl-CoA desaturases localized on the ER. Similar mixed function oxidases are also used to modify structural components of cells, hormones etc. so we will use the acyl-CoA desaturase as an example for this group of enzymes. In the acyl-CoA desaturase reaction molecular oxygen is used to oxidize both a fatty acid and NADH, each providing two of the the four electrons needed by the oxygen:

acyl-CoA desaturase reaction

The mammalian acyl desaturases are components in mini-electron transport systems on the surface of the endoplasmic reticulum, for example the Delta9-fatty acyl-CoA desaturase complex:

fatty acyl-CoA desaturase complex ETS diagram

Regulation of Fatty Acid Metabolism

Fatty acid metabolism is regulated both hormonally and via feed-back inhibition and feed-forward activation. Thus mobilization of free fatty acids from the adipose tissue results from low insulin levels. The free fatty acids are then transported through the blood to the rest of the body including the liver. In the liver fatty acid oxidation and ketone body synthesis is activated by glucagon. Note that glucagon and insulin levels are opposite: high insulin =low glucagon and vice-versa. So for low insulin will also have high glucagon, thus fatty acids will be released from the adipose and will be converted in the liver into ketone bodies.

The regulation of fatty acid oxidation, fatty acid synthesis and ketone body synthesis in the liver is summarized in the figure:

Regulation of Fatty Acid Metabolism in Liver diagram

Note that a LACK of insulin results in a release of fatty acids from adipose.

Photosynthesis

Look at chloroplast structure [Figure 15.16a & b, p 459; slide & below].

labeled chloroplast diagram

Chloroplast ultrastructure: 1) outer membrane, 2) intermembrane space, 3) inner membrane (1+2+3: envelope), 4) stroma (aqueous fluid), 5) thylakoid lumen (inside of thylakoid), 6) thylakoid membrane, 7) granum (stack of thylakoids), 8) thylakoid (lamella), 9) starch, 10) ribosome, 11) plastidial DNA, 12) plastoglobule (drop of lipids)

3-D cut-away model of chloroplast

Note how the thylacoids are stacked, with granal lamellae between stacked thylacoids, and stromal lamellae facing the plastisol (chloroplast matrix) [Figure 15.16, p 459]. The components of the photosynthetic ETS will be differentially distributed between these two lamellae [Figure 15.19, p 460]. Most of protons will be pumped into the lumen of the thylacoid system. (The important thing here is that the protons are being pumped out of the plastisol.) As in mitochondria, can use proton (Chemiosmotic) gradient to make ATP [Figure 15.18, p 460].

The Light Reactions

(Z Scheme)

Let's look a bit at chlorophyll and other pigments [Figure 15.1, p444] and its interaction with light.

Chlorophyll a strtuctural diagram

Chlorophyll a

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Different chlorophylls absorb light in different parts of the spectra as seen in the specta below for a and b [Figure 15.1, p 445]:

chlorophyll absrption spectra

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Find that there are many chlorophylls/active center. That is, only a small portion of the chlorophyll pool is actually involved in using the light energy (1/300 in Chlorella). What do the rest do? Act together as an antenna, transferring the captured photon energy via resonance energy transfer .[Figure 15.3, p 446]. The transfers between molecules take <10-10 sec with an efficiency of >90%. Higher plants also have beta-carotenes [Figure 15.4, p 447] to absorb other light frequencies. Aquatic plants have different dyes, since only blue-green light penetrates to depth.

beta carotene structural diagram

beta carotene

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So what is light energy used for? Look at the "Z" scheme for photosynthetic electron transport. [Figure 15.14, p 456]

Z-scheme cartoon diagram

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z-scheme diagram

Note the three major complexes are not directly connected. Like the mitochondrial complexes they are connected by carriers which diffuse between them [Figure 15.11, p 453].

z-scheme complexes in membrane

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The cytochrome b6/cytochrome f complex is analogous to Complex III in mitochondria (Lecture 28, 10 April): same electron path and a (plastoquinone) Q cycle for proton pumping.

 

Q-cycle diagram

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The initial removal of electrons from water to give oxygen uses a manganese complex, proposed to be a cubane-like structure, or more recently, the structure below:

manganese cluster structural diafgram

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Note the stoichiometry: one O2 : 2 H2O : 8 hgreek letter nu : 4 e- : 8 H+ pumped : 3 ATP : 2 NADPH

2 H2O + 2 NADP+ + 3 ADP + 3 Pi + 8 hgreek letter nu right arrow 2 NADPH + 2 H+ + 3 ATP + O2

The overall proton pumping and ATP synthesis process is summarized in the figure showing the distribution of the components of the Z scheme above.

Cyclic photophosphorylation involves PSI and cytochrome b5f and the Q pool [Figure 15.14, p 456]. Cyclic stoichiometry = 4 hgreek letter nu : 8 H+ pumped : 2 ATP

The Dark Reactions

(Calvin Cycle)

We have just looked at the so-called Light Reactions of photosynthesis: the process whereby the energy in sunlight is converted by plants into biological useful energy (ATP) and reducing equivalents (NADPH). Now we want to look at how plants use ATP and NADPH to capture ("fix") carbon dioxide and make glucose: the Calvin Cycle.

Most of the reactions of the Calvin Cycle are familiar. Thus on the Calvin Cycle pathway diagram reactions 1-5 (numbered in bold italics) are from Gluconeogenesis/Glycolysis, and reactions 6-8 (numbered in outline font) are from the Pentose Phosphate Shunt, while reactions 9 & 11 are variations of familiar enzyme catalyzed reactions. The only really new reaction is catalyzed by Ribulose-1,5-bis phosphate carboxylase.

Let's begin our look at the Calvin cycle with the first unique reaction of the cycle: the phosphorylation of Ribulose-5-P to Ru-1,5-bis P by Ru-5-P kinase (reaction 9 on the pathway diagram). The strategy here is to create a precursor which is symmetrically phosphorylated so that it can be cleaved into two 3-PGA's.

In the next reaction, catalyzed by Ribulose-1,5-bis phosphate carboxylase (RuBisCo), carbon dioxide is added to the keto-carbon, giving a six carbon sugar which can now be cleaved into the two three carbon PGA's. As shown in the figure, RuBisCo catalyses the addition of a carbon dioxide to ribulose-1,5-bis phosphate, which then splits to give two molecules of 3-PGA (note the use of carbon dioxide NOT bicarbonate as we have seen in the past):

structural formulas showing the mechn ism of the RuBisCo reaction

Note that in the rearrangement resulting with the carboxylation, the final three carbons become essentially a PGA, in particular the keto carbon now has the same oxidation state as an acid carbon (recall that an alcohol elimination involves no formal oxidation, thus a C-C bond is formally the same oxidation as a C-OH bond). Of course the upper portion also now looks about like a PGA too. The attack by water and subsequent cleavage releases the first 3-PGA, leaving a carbanion on the enzyme. The enzyme constrains the addition of a proton to this achiral intermediate to give only the 3-PGA of the proper chirality.

FYI - RUBISCO Structure

The enzyme itself is large (MW= 560,000 daltons), consisting of 8 large (MW= 56,000) catalytic subunits and 8 small (MW= 14,000) regulatory subunits. The eight catalytic subunits form the core of the enzyme, with the interfaces between them forming eight catalytic sites. The enzyme requires magnesium for activity, and there is a copper in each oligomer.

 

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Last modified 24 April 2013