1.

The Glycolytic Pathway

A.
B.

Historical Perspective
Pathway Overview
The Reactions of Glycolysis

A.
B.
C.

Hexokinase: First ATP Utilization

Phosphoglucose Isomerase
Phosphofructokinase: Second ATP Utilization
-D. Aldolase
'E. Triose Phosphate Isomerase
F.
Glyceraldehyde-3-phosphate Dehydrogenase: First "High-Energy"
Intermediate Formation
G.Phosphoglycerate Kinase: First ATP Generation
H.Phosphoglycerate Mutase
I.
Enolase: Second "High-Energy" Intermediate Formatlon
J. Pyruvate

Klnase: Second ATP Generation

3. Fermentation: The Anaeroblc Fate of Pyruvate

A.
B.
C.

Homolactic Fermentation
Alcoholic Fermentation
Energetics of Fermentation

4 Control of Metabolic Flux

A.
B.

Flux Generation 1Control of Glycolysis in Muscle

Metabolism of Hexoses Other Than Glucose A. Fructose
B Galactose
C Mannose

At this point we commence our discussions of specific metabolic pathways by

considering glycolysis (Greek: glykos, sweet; lysis, loosening), the pathway by
which glucose is converted via fructose-1,6-bisphosphate to pyruvate with the
generation of 2 mol of ATP/mol of glucose. This sequence of 10 enzymatic
reactions, which is probably the most completely understood biochemi cal
pathway, plays a key role in energy metabolism by providing a significant
portion of the energy utilized by most organisms and by preparing glucose, as
well as other carbohydrates, for oxidative degradation.
In our study of glycolysis, and indeed of all of metabo lism, we shall attempt to
understand the pathway on four levels:

1.

The chemical interconversion steps, that is, the sequence of reactions by

which glucose is converted to the pathway's end products.

2.

The mechanism of the enzymatic conversion of each pathway intermediate to

its successor.

3.

The energetics of the conversions.

4.

The mechanisms controlling the flux (rate of flow) of metabolites through the
pathway.

The flux of metabolites through a pathway is remark ably sensitive to the

needs of the organism for the products of the pathway. Through an exquisitely
complex network of control mechanisms, flux through a path waris only as
great as required.
-------------------------------------------------------------

Section 16-1. The Glycolytic Pathway 1. THE GLYCOLYTIC PATHWAY

An overview of glucose metabolism is diagrammed in Fig. 16-1. Under ae robic
conditions, the pyruvate formed by glycolysis is further oxidized by the citric acid
cycle (Chap-ter 19) and oxidative phosphorylation (Chapter 20) to CO, and water.
Under anaerobic conditions, however, the pyru-vate is instead converted to a
reduced end product, which is lactate in muscle (homolactic fermentation; a
fermenta-tion is an anaerobic biological reaction process) and ethanol + CO, in
yeast (alcoholic fermentation). A. Historical Perspective The fermentation of glucose
to ethanol and CO2 by yeast (Fig. 16-2) has been a useful process since before the
dawn of recorded history. Winemaking and baking both exploit this process. Yet, the
scientific investigation of the mechanism of glycolysis began only in the latter half
of the nineteenth century. In the years 1854 to 1864, Louis Pasteur established
Glycolysis
Glucose 2ADP 2NAD+ + 2P, Fructose-1,6- bisphosphate 2ATP 2NADH
2Pyruvate
Oxidative phosphorylation
Figure 16-1 Glycolysis converts glucose to pyruvate while generating two ATPs.
Under anaerobic conditions, alcoholic fermentation of pyruvate occurs in yeast while
homolactic fermentation occurs in musde. Under aerobic conditions, pyruvate is
oxidized to H20 and CO2 via the citric acid cycle (Chapter 19) and oxidative
phosphorylation (Chapter 20).
Figure 16-2 An electron micrograph of yeast cells. [Biophoto AssO4

that fermentation is caused by microorganisms*;',,;- - not until 1897, however, that

Eduard Buchner strated that cell-free yeast extracts can also carry process. This
discovery refuted the then widel belief that fermentation, and every other
biologic4 cess, was mediated by some "vital force" inher living matter, and thereby
brought glycolysis wit province of chemistry. This was a major step development of
biochemistry as a science. Althou principle, the use of cell-free extracts enabled a
sy,s atic "dissection" of the reactions involved in the` way, the complete elucidation
of the glycolytic was still a long-range project because analytical niques for the
isolation and identification of diates and enzymes had to be developed concud- tg.ly
In the years 1905 to 1910, Arthur Harden and W Young made two important
discoveries: 1. Inorganic phosphate is required for fermentatio is incorporated into
fructose-.1,6-bisphospha intermediate in the process. 2. A cell-free yeast extract can
be separated, by di, into two fractions that are both required for f tation: a
nondialyzable heat-labile fraction named zymase; and a dialyzable, heat-stable tion
they called cozymase. It was later sho others that zymase is a mixture of enzymes
anct cozymase is a mixture of cofactors: coenzyrn as NAD+, ATP, and ADP, as well
as metal ioris
jn their efforts to identify pathway intermediates, the eady investigators of
glycolysis developed a general technique of metabolic investigation that is still
widely used today: Reagents, are found that inhibit the production o fpathway
products thereby causing the buildup of metabo-lites that can thett be identified as
pathway intermediates. ver years of investigafion that attempted to identify
glycolytic intermediates, various reagents were found at inhibit the production of
ethanol from glucose in east extracts The use of different inhibitors resuits in e
accumubtion of different intermecliates. For exarn-le, the addition of iodoacetate to
fermenting yeast ex-acts causes the buildpp of fructose-1,6-bisphosphate hile
addition of fluoride ion induces the accumulation 2-phosphoglycerate and 3phosphoglycerate. 0 0 0 0 1 HC-2 OH OPOr HCOPOr HC-3 OH 1 3Phosphoglycerate 2-Phosphoglycerate e rnechanisms by which these inhibitors act
are dis-,cussed in Sections 16-2F and 16-21, respectively. One remarkable finding of
these studies was that the intermediates and enzyme activities could be iso-not
only from yeast, but from a great variety of er organisms. With few exceptions (see
Problem 10), ng things all metabolize glucose by identical pathways. spite of their
enormous diversity, they share a common hemistry. y 1940, the efforts of many
investigators had come Iruition with the elucidation of the complete pathway
glycolysis. Three of these individuals, Gustav Emb-Otto Meyerhof, and Jacob Parnas,
have been hon-in that glycolysis is alternatively known as the bden-Meyerhof
-Parnas pathway. Other major "butors to the elucidation of this pathway were and
Gerti Cori, Carl Neuberg, Robert Robison, and Warburg. Pathway Overview ore
beginning our detailed discussion of the en-es of glycolysis let us first take a
moment to survey overall pathway as it fits in with animal metabolism Whole.
Glucose usually arises in the blood as a result e breakdown of higher
polysaccharides (Sections B, 10-2C, and 17-1), or from ats synthesis from

rbohydrate sources (gluconeogenesis; Section ) The fate of nonglucose hexoses is

discussed in on 16-5. Glucose enters most cells by a specific r that transports it
from the exterior of the cell into cytosol (Section 18-2). The enzymes of glycolysis
are ted in the cytosol, where they are only loosely assoc:hapfer 16. Glycolysis 427
ciated, if at all, with cell structures such as membranes, and apparently form no
organized complexes with each other. Glycolysis converts glucose to two C3 units
(pyruvate) of lower free energy in a process that harnesses the released free energy
to synthesize ATP from ADP and Pi. This pro-cess requires a pathway of chemically
coupled phosphoryl-transfer reactions (Sections 15-4 and 15-6). Thus the chemical
strategy of glycolysis is 1. Add phosphoryl groups to the glucose. 2. Chemically
convert phosphorylated intermediates into compounds with high phosphate grouptransfer potentials. 3. Chemically couple the subsequent hydrolysis of reactive
substances to ATP synthesis. The 10 enzyme-catalyzed reactions of glycolysis are
diagrammed in Fig. 16-3. Note that ATP is used early in the pathway to synthesize
phosphoryl compounds (Re-actions 1 and 3) but is later resynthesized (Reactions 7
and 10). Glycolysis may therefore be considered to occur in two stages: Stage I
(ReaCtions 1-5): A preparatory stage in which the hexose glucose is phosphorylated
and cleaved to yield two molecules of the triose glyceraldehyde-3-phosphate. This
process utilizes two ATPs in a kind of energy investment.
Stage II (Reactions 6 -10): The two molecules of gly-ceraldehyde-3-phosphate are
converted to pyruvate with concomitant generation of four ATPs. Glycolysis
therefore has a net "profit" of two ATPs per glucose: Stage I consumes two ATPs;
Stage II produces four ATPs.
The overall reaction is Glucose + 2NAD+ + 2ADP + --> 2NADH + 2pyruvate + 2ATP
+ 2H20 + 411+ The enzymes of glycolysis, their subunit compositions, molecular
masses, and cofactors, are listed in Table 16-1. The Oxidizing Power of NAD+ Must
Be Recycled NAD+ is the primary oxidizing agent of glycolysis. The NADH produced
by this process (Fig. 16-3, Reac-tion 6) must be continually reoxidized to keep the
path-way supplied with NAD+. There are three common ways that this occurs (Fig.
16-1, bottom): 1. Under anaerobic conditions in muscle, NAD is re-generated when
NADH reduces pyruvate to lactate (homolactic fermentation; Section 16-3A). 2.
Under anaerobic conditions in yeast, pyruvate is de-carboxylated to acetaldehyde,
which is then reduced