Download Enzyme Mechanisms in Temperature and Pressure Adaptation of Off

A M . ZOOLOGIST, 11:425-485
(1971).
Enzyme Mechanisms in Temperature and Pressure Adaptation of
Off-Shore Benthic Organisms: The Basic Problem
PETER W. HOCHACHKA
Department of Zoology, University of British Columbia,
Vancouver 8, B. C, Canada
SYNOPSIS. The degree of participation of any given enzyme reaction in metabolism is
not determined by energy-volume relationships, but rather by enzyme affinities for key
regulatory ligands. In temperature adaptation, enzyme-substrate affinities often decrease
as thermal kinetic energy increases, thus compensating for temperature-induced
changes in reaction velocity at low substrate concentrations. Since the effects of
pressure are not unidirectional (some enzymes are accelerated; some are unaffected;
some are decelerated), a more functional solution in pressure adaptation is to
elaborate enzymes whose affinities for key metabolites are pressure independent. In
consequence, at physiological substrate concentrations, the pressure sensitivities of
catalysis and control of catalysis are probably held at a minimum.
restricted and often must occur at the molecular, rather than the whole-animal,
When an organism encounters an al- level of organization. A moment's contemtered environment, its metabolic response plation will make this distinction clear. An
to that change is "adaptive" if in proper artic marine mammal, for example, cosequence it activates an appropriate set of ordinates a whole series of physiological
reactions for answering the cell's new car- processes in order to maintain a constant
bon and/or energy demands. For example: body temperature in the face of the ex(1) the glucocorticoid-controlled produc- treme outer one, while, in contrast, an artion of glucose during periods of fasting, tic fish, because it cannot regulate its body
but not during satiated periods; (2) the temperature, must accept the environmenactivation of lipolysis in adipose tissues tal one as given. Here, temperature is imduring thermogenesis, but not during peri- pinging directly upon enzyme reaction
ods of thermal balance; (3) the activation rates and the organism's adaptive responses
of the urea cycle enzymes in Amphibian must begin at this point. The same is true
liver during terrestrial phases of life but for the effects of pressure on organisms in
not during aquatic larval stages. All such the water column.
categories of metabolic response can be
Biologists have known for many years
said to be patently adaptive. Formally, the that the metabolism of many poikilosignal and the precise response are the two therms is carefully adjusted to the thermal
components of the system that differ from regime in which they live. To choose an
example to example, but in each example extreme example, rates of energy metabowe are basically dealing with a network of lism, swimming performance, and growth
reactions whose degree of participation in of antartic fishes are as high at near 0°C as
metabolism at any time is automatically those of temperate zone fishes at 15-25°C
and usually autocatalytically controlled. By (Wohlschlag, 1961; Somero et al., 1968).
means of such control systems, most orga- This extreme cold adaptation of metabolic
nisms gain a certain degree of indepen- processes presumably requires evolutiondence from the outer environment. In ary time and has never been observed durwarm blooded forms this is manifest as ing shorter term adjustments, such as occur
homeothermy; in cold blooded forms, the during seasonal acclimatization (or acclidegree of independence from such parame- mation) of eurythermal temperate zone
ters as temperature or pressure is more fishes. Although the magnitude of the acINTRODUCTION
425
426
PETER W. HOCHACHKA
en
O
FIG. 1. Rates o£ O2 uptake by minced tuna red
muscle (T-RM), tuna white muscle (T-WM) and
skeletal muscle from the white rat (Rat-M). Data
redrawn from Cordon (19G8).
Rates of CO,
production from aceta te-1-C11 oxidation by trout
liver slices at different temperatures are shown in
the middle curve (x symbols). O2 uptake is in
mm'/gm/hour. C"O2 production is in picocuries/mg protein/hour. Data redrawn from
Dean. (1969).
climation response is less marked, it is in
the same direction; thus, the respiration of
tissues from cold acclimated fish is usually
higher than that of warm acclimated ones
at all temperatures examined (see Precht,
1968; Fry and Hochachka, 1970). Finally,
a number of examples of temperature independent metabolic processes have been
discovered in poikilothermic organisms,
and these represent another category of
thermal compensation. Glycogen synthesis
in insects can be temperature-independent
over quite broad thermal ranges (van
Handel, 1966); O2 uptake by intertidal
organisms can display Q l n values of near
unity (Newell, 1966); the respiration rate
of tuna muscle minces (Gordon, 1968) and
the oxidation of acetate-1-C14 by salmonid tissues (Dean, 1969) are stable over
20-30°C ranges (Fig. 1). Such immediate
thermal compensation is surely as dramatic
as the extreme cold adaptation of metabolism seen in the antarctic fishes.
For convenience we can distinguish
three time-courses of temperature adaptation in poikilotherms: (1) evolutionary
adaptation, (2) thermal acclimation, and
(3) immediate thermal compensation.
Comparable time-courses of exposure to
high and/or variable pressures are probably encountered by organisms in the marine water column. However, studies on the
effects of long-term exposures to high
piessutt's are not abundant, mainly be-
427
THE BASIC PROBLEM
u
o
2 -
330
353
5
10 /
FIG. 2. Arrhenius plot of liver citrate synthase activities from cold (2°C) acclimated trout at saturating oxaloacetate and acetylCoA concentrations is
shown in the curve labeled Vmax. The calculated
activation energy is 8.8 kcal/mole, corresponding to
a Qio of about 1.7. In the two lower curves,
oxaloacetate concentrations were at saturating
levels, but acetylCoA levels were 0.01 and 0.023
raM as shown. It is evident that the Qlo is dramatically decreased at low substrate concentrations.
Data redrawn from Hochachka and Lewis (1970).
Similar data are available for several different enzyme systems and are discussed by Hochachka and
Somero (1971).
cause the requisite technology is unavailable. For this reason, I will restrict this
paper to immediate effects of temperature
in comparison with the effects of pressure
upon enzymes of deep-sea organisms.
given in °K. A typical graphical presentation of this equation is shown in Figure 2
(for curve labeled, Vmad;). For workers interested in temperature adaptation, two
portions of the Arrhenius curve are of interest—the activation energy which is directly proportional to the slope of the line,
and the thermal optimum. Earlier workers
considered that either characteristic or both
should correlate with the habitat temperature, but as more data became available, it
became evident that neither does (see Hochachka and Somero, 1971). In regard to
thermal optima, the argument is simply
that for most enzymes these are far beyond
the lethal limits for the organism. As regards the value of /x, the situation is somewhat less clear. Basically, the idea commonly held is that selection would favor en-
IMMEDIATE EFFECTS OF TEMPERATURE
ON POIKILOTHERMIC ENZYMES
Classically, the effect of temperature on
enzyme reaction rates was interpreted according to the Arrhenius relationship given by equation (1):
* = 2.3R
log KTi — log KT(>
1
i —
1
(1)
o
where ^ is the activation energy, R the gas
constant, K the rate constant for the reaction at 2 different temperatures, T1 and To,
428
PETER W. HOCHACHKA
CITRATE SYNTHASE
1
-
FIG. 3. Effect of temperature on the Km of acetylCoA for liver citrate synthase from cold acclimated rainbow trout. Data redrawn from Hochachka and Lewis (1970). As temperature in-
creases, the apparent enzyme-substrate affinity (reciprocal of the Km) decreases, thus compensating
the reaction for changes in the thermal energy of
the reactants.
zymes of high catalytic efficiency (and
hence low ji value) in organisms living at
low temperatures. Where JX is an acceptable
measure of catalytic efficiency, the correlation may indeed hold. But the turnover
number of an enzyme can also be strongly
influenced by activation entropy, which has
been largely ignored by earlier studies. This
may account for the absence of an obvious
correlation between the ^ value for any
two homologous enzymes and the thermal
habitat of the parent species, whereas such
a correlation does appear to exist for the
turnover number of enzymes. (See Hochachka and Somero, 1971, for a further
discussion of this point.)
The situation, however, is still more
complex. As it> e\ident in Figure 2, the
slope of Arrhenius plots can vary dramatically depending upon substrate concentra-
tions. At low substrate levels, the temperature coefficient (Qlo) is dramatically reduced and can take on values of less than
1.0. The explanation for such seemingly
anomalous thermal properties is to be
found in the nature of the catalyst. For a
large number of enzyme systems, we have
found that apparent Michaelis constant
(Km) varies directly with temperature over
at least a part of the biological range.
Over this range, as temperature increases,
the apparent enzyme-substrate affinity decreases in such a way as to compensate for
the thermal energy change (Fig. 3). Thus,
under low (physiological) substrate concentrations, the reaction rate is being determined by the kinetic properties of the
catalyst rather than thermudyniunit parameters (Hochachka and Somero, 1971).
In many wa)s, there is a strong similarity
T H E BASIC PROBLEM
429
FIG. 4. Comparison of the effects of the positive
modulator, AMP, and of low temperatures upon
the activity of king crab phosphofructokinase. Data
redrawn from Freed (1971).
between the effects of temperature and
the effects of metabolite modulators on
enzyme-substrate affinities. A particularly clear example is to be found in the
case of king crab phosphofructokinase,
where the effects of low temperature and
of the positive modulator, AMP, are strikingly comparable (Freed, 1971) (Fig. 4).
In this example, as in the case of many
regulatory enzymes, catalytic activity is regulated by the binding of specific metabolites at sites separate from the catalytic
site; usually, these modulators alter the apparent enzyme-substrate affinity with no
necessary effect on the maximal velocity
(Vmaa:) of the reaction. Positive modulators
decrease the apparent Km, and sometimes
convert the sigmoidal curve into a hyperbolic one. (At low substrate levels, this is
tantamount to activating the enzyme.) In
both these effects, the action of positive
modulators is analogous to that of low
temperature. Somero (1969) also has described an elegant example of this in the
case of king crab pyruvate kinase (PK):
At low temperatures, the apparent Km of
the substrate (PEP) is reduced, and substrate saturation curves become hyperbolic
rather than sigmoidal.
Another fundamental characteristic of
regulatory enzymes is that saturation curves for positive or negative modulators also
are often sigmoidal. An important implication of this is that small changes in modulator concentration can lead to relatively
large changes in the activity of the enzyme.
This property coupled with others, such
as product activation, leads to regulatory
behavior which approximates an "on-off"
switch. In general, "on-off" switch mechanisms of poikilothermic enzymes seem to be
less temperature sensitive than are enzyme-
430
PETER W. HOCHACHKA
substrate interactions (see Hocliachka and
Somero, 1971). This admirably suits poikilothermic existence, for the regulation of
enzyme activity can then be achieved with
equal efficiency over the entire biological
temperature range, irrespective of the
effects of temperature on maximum velocities.
Because of the fundamental role of Km
modulation in enzyme regulation in general, then, it is not too surprising that the
effects of change in the thermal energy of
reactants can be counteracted by appropriate and instantaneous effects of temperature, presumably upon enzyme conformation and hence upon enzyme-substrate
affinities. But, such actions of temperature
are complicated in deep-sea fishes by continuous high pressures and in midwater
fishes by linear changes in pressure encountered during vertical migrations
through the water column. Hence, it is
informative to examine the effects of pressure on enzymes in the context discussed
above.
IMMEDIATE EFFECTS OF PRESSURE
ON ENZYME REACTION RATES
The basic action of pressure upon various metabolic processes is fairly well
documented. Nearly 40 years ago, Johnson
and Eyring and coworkers described the
fundamental theorem relating pressure
and reaction velocities (see Johnson and
Eyring, 1970). This relationship is given in
equation (2):
AV*=2.3RT
(2)
where AJ7* is the volume change of activation, R the gas constant, K the velocity
constant at pressure px and p2. This equation for the relation between pressure and
reaction rate is of the same form as the
Arrhenius equation for the relation between temperatures and rate. According to
the Arrhenius theory of absolute rates, it
is necessary to assume the existence of an
activated complex to account for the large
effect of temperature on reaction rates. In-
deed, the reaction rate can be taken as a
measure of the concentration of the activated complex. So long as the activated
complex represents a more energy-rich
configuration than the average value of its
constituents, a rise in temperature will increase the rate. Where this is no longer
true, the rate process goes through a maximum.
A comparable view holds with regard to
pressure. Here, A P can be taken as the
ratio
volume of the activated complex
volume of the reactants
So long as the volume of the activated
complex exceeds the average volume of its
constituents outside the complex, pressure
retards the reaction. At the point where
the volumes become equal, there is no
change in the reaction rate under pressure.
When the volume of the activated complex is less than that of the reactants, pressure accelerates the reaction rate (Fig. 5).
In this latter case, the reaction rate can go
through a pressure optimum, the actual
pressure at which the optimum occurs often depending upon the temperature (Fig.
5). Most previous workers in this field
have obtained results comparable to those
shown in Figure 5 (see Morita, 1967;
ZoBell, 1970; Johnson and Eyring, 1970)
and this seems true even at higher levels of
organization. (See Brown et ah, 1958, for
effects of pressure on tension developed in
turtle heart at different temperatures.)
The above noted interaction between
temperature and pressure is an important
aspect of this problem but one which often
has been given inappropriate consideration. ZoBell and Hittle (1969) for example found that the rate of starch hydrolysis
by more than a hundred species of marine
bacteria is roughly proportional to their
rate of growth at different pressures and
temperatures. To gain further information
on this matter, these workers then examined a partially purified a-amylase prepared from Bacillus sitbtilis, a well studied non-marine bacterium! This enzyme was found to be accelerated by
"deep-sea pressures" at temperatures be-
T H E BASIC PROBLEM
431
PRESSURE
FIG. 5. Steady state levels o£ luminescence intensity
in cell free extracts of the luminous bacterium
Achromobacter fishcheri. Data redrawn from
Strehler and Johnson (1954). See also Johnson and
Eyring (1970) for recent discussion of this
system.
tween 25-60°C, but not at lower temperatures {see also ZoBell, 1970). The authors
point out that such acceleration is predicted by theory since starch hydrolysis is a
reaction that takes place with a volume
decrease. But starch hydrolysis in marine
bottom bacteria occurs at 2-4°, where no
pressure activation was noted; hence, either the data or the theory are inadequate.
The authors further point out that,
whereas "deep-sea pressures" inactivate the
enzyme at temperatures below 35°C, they
stabilize it at temperatures between 4560°C. The implication is that such pressure-stabilization is of some biological significance. Since nowhere on this planet do
temperatures of the ocean bottom ever
come close to 45°C, their results indicate
(1) that the enzyme characteristics measured are biologically meaningless, or (2)
that the homologous enzyme from ocean
bottom bacteria must display entirely different properties—or both. This field is riddled with examples of a similar kind (see
review by Morita, 1967). Even Schleiper
(1968), who is quite aware of this problem, concluded that because the pressure
resistance of tissue of Mytilus eclulis is
greater at 20° than at 10°, deep-sea animals would cope more successfully with
ocean bottom pressures at higher temperatures. Possibly this is true, but certainly it
is biologically irrelevant. The point is that
temperature of the ocean bottom is
2-4°C; this is where off-shore benthic organisms live, and within the group this is
where adaptation to pressure, if it is going
to occur, must occur.
Much of the available literature also
stresses the above formal analogies between the equations relating pressure and
temperature to reaction rates of various
metabolic processes (equations 1 & 2
above). However, I think that it is important to stress the following difference.
Whereas temperature affects all chemical reactions in the same way (by altering
the kinetic energy of the reactants), pres-
432
PETER W. HOCHACHKA
G1P
Si- 1 - 7
-3.4
Hk
-
5.0
G6P '•••"•"••|ii'"""iiti1111"11"1*1* 6PG
GLUCOSE
G6PDH
G6 Pase
- 4.0
2:
§5
si
i
F6P
FIG. 6. The G6P crossroads in metabolism. Each of
the enzymes involved directly in G6P production or utilization is shown in the Figure along
with the standard free energy change (AG°) which
accompanies each reaction. Phosphoglucomutase
(PGM); hexokinase (Hk); glucose-6-phosphatase
(G6Pase); hexose isomerase (isomerase); and glucoses-phosphate dehydrogenase (G6PDH) are the
abbreviations used. Three of these enzymes (Hk,
G6Pase, and G6PDH) are known to be involved in
the partitioning of G6P between these various reaction pathways.
sure activates some, retards some, and does
not affect others. What is more, pressure
can bring about all three of these effects on
a given enzyme-catalyzed reaction depending upon the temperature, or more precisely, upon the enzyme conformation
adopted at different temperatures. It is evident, therefore, that from a functional and
evolutionary point of view, pressure is an
entirely different kind of physical parameter than is temperature. This conclusion
has some interesting implications.
Consider for example the effects of pressure on a branch point in metabolism such
as the G6P crossroads (Fig. 6). In theory,
AV* for each reaction could be calculated
from knowledge of the geometry of the initial and transition state of the reacting
components. In practice, of course, sufficient information about the configuration
of the transition state and its neighboring
molecules is a\ailable for only the simplest
of reactions, and is certainlv unavailable
for the reactions of G6P metabolism.
Hence, to predict the sign and magnitude
of AV* for such reactions approximations
have to be used. These usually assume
AV* to be the sum of two terms: the "structural" term, A\V*, is the change in volume
of die reacting molecules during bondbreakage and bond-formation; the "solvation" term, A^V*, is the accompanying
volume change due to interaction with surrounding molecules. When ionic species
are formed or disappear in the transition
state, the "solvation" contribution to AV*
usually excedes the "structural" volume
change in the reacting molecules. As a first
approximation, then, we may conclude that,
in the absence of regulation on the part of
the organism, the "charge type" of the reaction will determine the effect of pressure
upon its rate.
For a branchpoint such as the G6P crossroads (Fig. 6), the ionic species invohed in
THE BASIC PROBLEM
each reaction as well as the reaction mechanisms are specific to each competing pathway. In addition to different "solvation"
contributions to AJ7* for each of the G6P
reactions, each is catalyzed by different
kinds of enzymes (some are membrane
bound; some are apparently free in solution; some are single-subunit types; some
are oligomeric). For these reasons, the effects of pressure upon each of these reactions will be quantitatively different. Now
three of the enzymes (Hexokinase, G6Pase,
G6P dehydrogenase) undoubtedly are involved in regulation of G6P metabolism in
liver (Scrutton and Utter, 1968; Atkinson,
1966). Hence, even in such a simple case,
it is perfectly clear that the flow of carbon through metabolic branchpoints may
be profoundly affected by pressure changes
and therefore that the control requirements
at such points may depend critically upon
the pressure.
For the same reasons, the effects of pressure upon the partitioning of carbon and
energy between various metabolic pathways may be extreme. Some suggestive data
are available in the literature. Chumak
(1959) observed that in a marine bacterium both the uptake and the manner in
which glucose is metabolized vary dramatically with pressure: CO2 production from
glucose is inhibited by pressure, whereas
glucose uptake and the production of acid
end products both increase markedly. Similarly, the extensive studies of pressure
effects on developing marine eggs demonstrate that the partitioning of carbon and
energy between DNA synthesis, formation
of the mitotic spindle, chromosomal movements, and cytokinesis is markedly affected
by pressure (see Zimmerman, 1970) . But,
these studies and a host of others of their
type (see ZoBell, 1970; Morita, 1967) were
designed more from an empirical than
from a functional point of view; hence,
interpretation of the data in mechanistic
terms is impossible. In any event, it is clear
that on theoretical grounds, basic problems of integrating various effects of pressure suggest essential requirements for
pressure adaptation at the molecular level.
433
The wide abundance of benthic and
mesopelagic organisms which thrive under
high and/or variable pressures indicate
that the enzymatic problems imposed by
this parameter are circumvented in nature.
The question is how. Most earlier studies
of pressure effects on enzymes (often using
enzymes from mammalian and nonmarine bacterial sources) have measured
pressure effects on catalysis under conditions of saturating substrates, cofactors,
and/or coenzymes. And, indeed, the formal
analogies between the effects of temperature and pressure are valid only under
these conditions, because both the Arrhenius and the Johnson-Eyring equations
(equations 1 and 2 above) assume a saturated catalyst. Unfortunately, it is now almost axiomatic in the literature on metabolic control that in vivo most enzymes
never "see" saturating concentrations of
substrates (Atkinson, 1969; Scrutton and
Utter, 1968). As already mentioned, under
in vivo conditions, regulation of enzyme
activity is usually achieved through the
control of enzyme affinities for unique ligands (substrates, cofactors, modulators).
Hence it is important to have information
on the effects of pressure on these parameters. In this context, our initial studies of
fructose diphosphatase (FDPase) in the
liver of deep-sea fish indicate that affinities
of the enzyme for its substrate (FDP), its
cofactor (Mg2+) and its negative modulator (AMP) are all independent of pressure
(Hochachka et al, 1970). This result is
particularly relevant from a functional
point of view, for it suggests that at low
substrate concentrations control of catalysis
is pressure independent, irrespective of
what pressure does to the maximum velocity. In the studies presented in this report, this general conclusion is fully
confirmed for several enzyme systems.
When the opportunity arose to extend
our studies on enzymes from deep-sea fish,
we chose a series of enzymes whose physiological functions and regulatory properties
arc fairly well documented. These enzymes,
the reactions they catalyze, and their
434
PETER W. HOCHACHKA
G6P
F6P
High ATP
Low AMP
1
TP
NAD—i
NADH
#
PEP
NADH
NAD
PYR
LAC
FIG. 7. A number of key steps and control circuits
in glycolysis. Phosphofructokinase (PFK), which
catalyzes a transphosphorylation of F6P thus forming FDP, is inhibited by energy-saturating conditions (high ATP) while the reverse reaction, catalyzed by fructose diphosphatase (FDPase), is simultaneously deinhibited. Under energy-depleted
conditions, high le\els of AMP inhibit FDPase and
concurrently activate PFK. The pyrmatc kinase
(PK) catalyzed conversion of PEP to pyruvate (pyr)
is likewise under stringent regulation by feedforward FDP activation and feedback ATP inhibition.
The comersion of pjr to lac (lactate), catalyzed by
lactate dchydrogenase isozymcs, can be involved in
the regulation of gHcolysis by affecting the supply
of NAD. The effects of pressure on FDPase, PKK,
PK, and I.DHs will be examined in the companion
papers.
integration into glycolysis and energy metabolism as a whole are diagrammed simply
in Figure 7. More detailed properties are
described in the companion papers. At this
point, what I want to stress is that in benthic fishes, whether
pressure
accelerates
THE BASIC PROBLEM
maximum velocities (as in the case of
phosphofructokinase and FDPase), does
not change them (as in the case of LDH)
or retards them (as in the case of PK),
enzyme-substrate and enzyme-modulator
affinities are largely insensitive to pressure.
These studies o£ temperature and pressure effects on poikilothermic enzymes lead
to a unifying hypothesis; namely, that at
physiological substrate concentrations enzyme reaction rates are controlled by key
kinetic parameters rather than by energyvolume relationships. In the case of temperature adaptation, enzyme-substrate affinities often are adjusted to compensate
for the unidirectional effect of thermal
change. In pressure adaptation, because the
effects of pressure are not unidirectional, a
more functional solution to the problem is
to maintain enzymes whose affinities for
their substrates are pressure-independent.
In consequence, at physiological substrate
concentrations, the pressure sensitivities of
catalysis and control of catalysis are probably held at a minimum. If, in regard to
pressure, these catalytic and regulatory
properties are widespread in enzymes ot
benthic and mesopelagic organisms, they
supply a sufficient mechanism for avoiding
uncontrollable effects of high pressure on
chemical reactions.
REFERENCES
Atkinson, D. E. 1966. Regulation of enzyme activity. Aiinu. Rev. Biochem. 35:85-124.
Atkinson, D. E. 1969. Limitation of metabolite
concentrations and the conservation of solvent
capacity in the living cell. Curr. Top. in Cell.
Regul. 1:29-43.
Brown, D. E. S., K. F. Cuthe, H. C. Lawler, and M.
P. Carpenter. 1958. The pressure, temperature and ion relations of myosin ATPase. J. Cell.
Comp. Physiol. 5259-77.
Chumak, M. D. 1959. The effect of high pressure
on the rate of glucose utili/.ation by barotolerant
bacteria. Dokl. Rlol. Sci. 126:524-526.
Dean, ]. M. 1969. The metabolism of tissues of
thermally acclimated trout (Salmo gairdneri).
Comp. Biochem. Physiol. 29:185-196.
Freed, J. M. 1971. Temperature effects on muscle
phosphofructokinase of the Alaskan king crab,
Paralithodes camtschatica Comp. Biochem.
Physiol. 39-765-774.
Fry, F. E. J., and P. W. Hochachka. 1970. Fish, p.
79-134. In C. Whittow [ed.], Comparative
435
physiology of thermoregulation. Academic Press,
New York.
Gordon, M. S. 1968. Oxygen consumption of red
and white muscles from tuna fishes. Science
159:87-90.
Hochachka, P. W., and J. K. Lewis. 1970. Enzyme
variants in thermal acclimation. Trout liver citrate synthases. J. Biol. Chem. 245:6567-6573.
Hochachka, P. W., D. E. Schneider, and A.
Kuznetsov. 1970. Interacting pressure and temperature effects on enzymes of marine poikilotherms: Catalytic and regulatory properties of
FDPase from deep and shallow-water fishes. Mar.
Biol. 7:285-293.
Hochachka, P. W., and G. N. Somero. 1971. Biochemical adaptation to the environment, p. 99156. In W. S. Hoar, and D. J. Randall [ed.], Fish
physiology, Vol. 6. Academic Press, New York. (In
press).
Johnson, F. H., and H. Eyring. 1970. The
kinetic basis of pressure effects in biology and
chemistry, p. 1-44. In A. M. Zimmerman [ed.],
High pressure effects on cellular processes.
Academic Press, New York.
Morita, R. Y. 1967. Effects of hydrostatic pressure
on marine microorganisms. Oceanogr. Mar. Biol.
Annu. Rev. 5:187-203j
Newell, R. C. 1966. Effect of temperature on the
metabolism of poikilotherms. Nature (London)
212:426-428.
Precht, H. 1968. Der Einfluss "normaler" temperaturen auf lebensprozesse bei wechselwarmen tierne unter ausschluss der wachstums- und entwicklungsprozesse. Helgolaender Wiss. Meeresunters. 18:487-548.
Schleiper, C. 1968. High pressure effects on marine invertegrates and fishes. Mar. Biol. 2:5-12.
Scrutton, M. C, and M. F. Utter. 1968. The regulation of glycolysis and gluconeogenesis in animal
tissues. Annu. Rev. Biochem. 37:249-302.
Somero, G. N. 1969. Pyruvate kinase variants of the
Alaska king crab: Evidence for a temperature-dependent interconversion between two
forms having distinct- and adaptive- kinetic
properties. Biochem. J. 114:237-241.
Somero, G. N., A. C. Giese, and D. E. Wohlschlag.
1968. Cold adaptation of the Antarctic fish Trematomus bernacchii. Comp. Biochem. Physiol.
26:223-233.
Strehler, B. L., and F. H. Johnson. 1954. The
temperature-pressure-inhibitor relations of bacterial luminescence in vitro. Proc. Nat. Acad. Sci.
U.S. 40:606-617.
van Handel, E. 1966. The thermal dependence of
the rates of glycogen and triglyceride synthesis in
the mosquito. J. Exp. Biol. 44:523-528.
Wohlschlag, D. E. 1961. Growth of an Antarctic
fish at freezing temperatures. Copcia 11-18.
Zimmerman, A. M. 1970. High pressure studies on
synthesis in marine eggs, p. 235-257. In A. M.
Zimmerman [ed.], High pressure effects on cellular processes. Academic Press, New York.
ZoBell, C. E. 1970. Pressure effects on morphology
and life processes of bacteria, p. 85-130. In A. M.
Zimmerman [ed.], High pressure effects on cellular processes. Academic Press, New York.
ZoBell, C. E., and L. L. Hittle. 1969. Deep-sea
pressure effects on starch hydrolysis by marine
bacteria. J. Oceanogr. Soc. Jap. 25:36-47.