Bariloche protein symposium argentine society for biochemistry and molecular biology

155
BIOCELL, 27 (Suppl. I), 2003
LI-P30.
A SPIN LABEL STUDY OF THE INTERACTION OF
BIOLOGICALLY ACTIVE TERPENES WITH MODEL
MEMBRANES
Turina, Anahi del V.
1
; Clop, Eduardo M.
1
; Perillo, Maria, A.
1
 and
Schreier, Shirley
2
1
Cátedra de Biofísica-Química, Facultad de Ciencias Exactas,
Físicas y Naturales, Universidad Nacional de Córdoba, Córdoba,
Argentina; 
2
Departamento de Bioquímica, Instituto de Química,
Universidade de São Paulo, São Paulo, Brasil. E-mail:
[e-mail]
Terpenes exert a variety of biological activities. We have
investigated the interaction between a series of terpenes (cineol,
geraniol, thymol, menthol, and camphor) and phospholipid (in the
absence and presence of 25 mole% cholesterol) model membranes
by means of spin labeling EPR. EPR spectra of incorporated lipid
spin probes (doxyl derivatives of stearic acid carrying the nitroxide
moiety at carbons 5, 5-SASL, and 12, 12-SASL) were obtained as
a function of terpene concentration. The spectra indicated that the
degree of molecular freedom of the probes increased with
increasing terpene concentration until a saturation state was
obtained. Saturation was ascribed to the achievement of the
solubility of the compounds in aqueous solution. It was found that
the different molecules affected molecular motion to a different
extent, probably due to the contribution of intrinsic differences in
chemical structure and to the degree of partitioning in the model
membranes. In addition, all terpenes except for camphor, formed
micelle-like aggregates at enough concentrations. At these
concentrations, in the presence of membranes, the compounds were
able to form mixed terpene:phospholipid micelles.
Supported by CONICET, SeCyT,ACC, Foncyt, FAPESP, CNPq.
LI-P31.
EFFECTS OF ISCHEMIA/REPERFUSION AND TRAUMA
ON TESTICULAR LIPIDS WITH VERY LONG CHAIN
POLYUNSATURATED FATTY ACIDS
Zanetti SR, Maldonado EN, and Aveldaño MI.
INIBIBB, CONICET-UNS, 8000 Bahía Blanca, Argentina. E-mail:
[e-mail]
Rat testicular lipids contain, besides molecular species with usual
fatty acids (FA), species with very long chain polyunsaturated fatty
acids (VLCPUFA). One of these lipids is ceramide (Cer). In search
for possible functions of VLCPUFA-containing lipids, we studied
them it in situations that in other tissues are known to result in
cellular stress-related changes. In testis, ischemia induced by
spermatic cord torsion followed by reperfusion, and trauma
provoked by puncture, resulted, 1-2 days afterwards, in Cer with
an increased proportion of FA other than VLCPUFA. These Cer
could be related with the activation of mechanisms responsible
for the selective death of germinal cells associated with both
damaging conditions. One of the early consequences of testicular
injury in both instances was seen in a neutral lipid not related to
Cer: the cholesterol esters (CE). In this lipid, 7 days after the
experimental harm, the proportion of VLCPUFA decreased while
that of normal PUFA (18:2n-6 to 22:5n-6) augmented. The changes
in the FA of Cer and CE occurred in the presence of virtually
unchanged triglycerides and phospholipids, the major testicular
lipids. The latter changed later on in the process, suggesting a
relation to the testicular atrophy eventually triggered by the present
conditions.
LI-P32.
AMINO ACID CHANGES IN THE INTERMEMBRANE-
SPACE DOMAIN OF MITOCHONDRIAL GLYCEROL-3-
PHOSPHATE ACYLTRANSFERASE ALTER  THE
CATALYTIC ACTIVITY  IN THE  CYTOSOLIC  N-
TERMINAL DOMAIN
Pellon Maison M
1
, Cunningham ML
1
, Coleman RA
2
, and Gonzalez-
Baró MR
1
.
1
INIBIOLP (CONICET-UNLP) and 
2
Department of Nutrition,
University of North Carolina, NC, USA.
Mitochondrial glycerol-3-phosphate acyltransferase (GPAT, 828-
aa) catalyses the first step in glycerolipid synthesis. It is inserted
in the outer membrane by two transmembrane domains, having
its active site located in the N-terminal domain, and the C-terminal
domain, both facing the cytosol, and a 82-aa loop in the
intermembrane space. Even though the loop and active site lay on
opposite faces of the membrane, the insertion of a HA epitope
(YPYDVPDYA) in the loop (construct HA496) completely
inactivated the enzyme. To determine whether the change in the
size (91 aa instead of 82) or the polarity of the loop were
responsible for the inactivation of the enzyme, two additional loop-
mutated GPATs, one with an insertion of 9 neutral aa (loop91)
and one with a 42-aa deletion in the loop (loop40), plus the full-
length protein with a Flag in the C-terminus (GFlag) were
transiently expressed in CHO cells. The level of expression of
these constructs was normalized by immunodetection. Loop91 and
Loop40 yielded partially active enzymes (8.3 and 2.3% of the
GFlag activity), whereas HA496 activity was ‘lower than the empty
vector. We postulate that the loop is required for activity; it may
contribute to the proper folding of the enzyme or it may be involved
in supramolecular regulatory interactions.
LI-P33.
CADMIUM AND LIPID METABOLISM IN PITUITARY
Calderoni AM
1
, Oliveros LB
1
, Anton R
1
, Jahn GA
3
, Mohamed F
2
and Giménez MS
1
.
1
Laboratorios de Química Biológica and 
2
Histología, Facultad
de Qca Bqca. y Farmacia. Universidad Nacional de San Luis,
5700 San Luis. 
3
IMBECU, CRICYT, Mendoza.
It is known that  exposure to 15 ppm of cadmium as CdCl
2 in the
drinking water  by eight weeks decrease  the secretion of prolactine
and growth hormone. from  the anterior pituitary lobe (APL) of
adult male Wistar rat.In this work we study the effect of cadmium
on dispersed cells of APL on the phospholipase D activity and
phosphoinositolbiphosphate (PIP2) synthesis  and its relation with
the level of serum prolactin secretion.Also we study the
histoarchitecture of APL of rats which drunk 15 ppm of cadmium
as CdCl
2 in the drinking water  by eight weeks.The PLD activity
was measured by the incorporation of 
[3H]-myristate into
phosphatidylethanol trough the transphophatidylation reaction.
PIP2 synthesis was determined by the incorporation of  
[3H]-myo
inositol. The results showed that cadmium cause the next changes
in relation to the control (without metal exposition): The activity
of Phospholipase D decreased as PIP2 synthesis did it. In APL a
significant capillary dilatation and a leucocitary and macrophagic
invasion were  observed in cadmium treated rats, compared with
those of control. All these changes could be related to the low
level of prolactine, since the prolactine secretion it is known to be
mediated by cellular signals involve phosphoinositide pathways.

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