- Type of paper
: Paper in book
Title: Specificity in RNA:protein interactions; the recognition
of Escherichia coli glutamine tRNA
- Authors:
- Rogers, M.J.
- Weygand-Đurašević, Ivana (57696)
- Schwob, Etienne
- Sherman, J.M.
- Rogers, K.C.
- Thomann, H.U.
- Sylvers, L.A.
- Jahn, M.
- Inokuchi, H.
- Ohtsuka, Eiko
- Soll, Dieter
- Editors
- Nierhaus, K.H.
- Subramanian, A.R.
- Erdman, V.A.
- Franceschi, F.
- Wittman-Liebold, B.
Publisher: Plenum press
Year: 1993
Pages: from 47 to 66
Number of references: 33
Language: engleski
Summary:
A variety of genetic, biochemical and structural studies havebeen used to
determine factors ensuring the accuracy ofrecognition by aminoacyl-tRNA
synthetases for tRNA. The identityelements of Escherichia coli tRNA(Gln)
are located mainly in theanticodon and acceptor stem, and ensure the
accurate recognitionof the tRNA by glutaminyl-tRNA synthetase. We summarize
a numberof experimental tehniques to define the accuracy ofaminoacylation
in vivo and in vitro.
Keywords: aminoacyl-tRNA synthetases, tRNA
- Type of paper
: Paper in book
Title: Crick, Francis Harry Compton
- Authors:
- Kućan, Željko (24372)
- Editors
- Vince, R.
Publisher: Biblioteka Hrvatski radio
Year: 1994
Pages: from 152 to 155
Language: hrvatski
- Type of paper
: Paper in book
Title: Watson, James Dewey
- Authors:
- Kućan, Željko (24372)
- Editors
- Vince, R.
Publisher: Biblioteka Hrvatski radio
Year: 1994
Pages: from 651 to 654
Language: hrvatski
- Type of paper
: Paper in book
Title: My early days at the Rockefeller
- Authors:
- Kućan, Željko (24372)
- Editors
- Paar, Vladimir
Publisher: Školska knjiga
Year: 1995
Pages: from 60 to 62
Language: engleski
- Type of paper
: Paper in journal
Title:
- Authors:
- Kućan, Željko (24372)
Journal: Filozofska istraĐivanja
Volume: 15
Year: 1995
Pages: from 93 to 101
Language: hrvatski
- Type of paper
: Paper in journal
Title:
- Authors:
- Kućan, Željko (24372)
Journal: Encyclopaedia Moderna
Volume: 14
Year: 1993
Pages: from 115 to 121
Language: hrvatski
- Type of paper
: Summary in proceedings
Title: Complexes of yeast tyrosil-tRNA synthethase with tRNA
- Authors:
- Rubelj, Ivica (108443)
- Weygand-Đurašević, Ivana (57696)
- Kućan, Željko (24372)
Proceedings title: 15th International Congress of Biochemistry
Language: engleski
Place: Jerusalem, Israel
Year: 1991
Pages: from 149 to 149
Meeting: 15th International Congress of Biochemistry
Held: from 08/04/91 to 08/08/91
Summary: Yeast tyrosyl-tRNA synthethase (TyrRS) is an alpha2 enzyme
ofthe subunit Mr=45000. Complexes of TyrRS with tRNA were studiedby
polyacrylamide gel elektrophoresis. In the absence of ATP, twoA2-enzymes
bind any cognate or noncognate tRNA molecule andprotect it efficiently
against nuclease digestion. Competitionexperiments show that in the
(alpha2)2-tRNA type of complex thebinding of tRNA(Tyr) is only 10-fold
stronger than the bindingof non-cognate tRNAs. However, even the binding of
non-cognatetRNA inhibits the pyrophosphate exchange rection on TyrRS.
Excessof cognate tRNA(Tyr) and/or ATP prevents the binding ofnon-cognate
tRNA and flavours the formation of alpha2-tRNA(Tyr)complex, in which the
tRNA is only partially shielded againstnucleases. ATP can be replaced by
GTP, a competetive inhibitor ofTyrRS, in the formation of the alpha2-tRNA
complex, but not inthe aminoacylation reaction. The alpha2-enzyme,
stabilized byATP, seems to be the only specific and catalytically active
form.On the other hand, the (alpha2)2-form, as well as the alphasubunit
alone, show very strong general affinity towards nucleicacids, but are
completely devoid of specificity towards tRNA(Tyr)
Keywords: tyrosyl-tRNA, electrophoresis, digestion,
- Type of paper
: Summary in proceedings
Title: The accurate recognition of Escherichia coli glutamine
tRNA
- Authors:
- Rogers, M.J.
- Weygand-Đurašević, Ivana (57696)
- Schwob, Etienne
- Sherman, J.M.
- Rogers, K.C.
- Thomann, H.U.
- Sylvers, L.A.
- Jahn, M.
- Fett, R.
- Inokuchi, H.
- Ohtsuka, Eiko
- Soll, Dieter
- Editors
- Blanquet, S.
Proceedings title: 15th International tRNA workshop
Language: engleski
Place: Cap d'Agde, Francuska
Year: 1993
Pages: from 144 to 144
Meeting: 15th International tRNA workshop
Held: from 05/30/93 to 06/04/93
Summary: Genetic, Biochemical and biophysical tehniques have been
used todefine the factors ensuring the accurate recognition by
E.coliglutaminyl-tRNA synthethase (GlnRS) for its substrates: ATP,glutamine
and tRNA(Gln). This has resulted in the first RNA:protein strukture
determined by X-ray crystallography (1,2). Thaaccuracy of amiloacylation of
the tRNA depends on two principles:competetion between synthetases in the
cell (3,4), and therecognition of identity elements principally located in
theanticodon and acceptor stem of tRNA(Gln) (5,6). The recognitionof
individual nucleotides, as in interaction with the 2-aminogroup of G, is
also apparent from studies withinosine-supstituted tRNAs (7). A genetic
selection has beenfruitful in obtaining mutants of GlnRS which confer
altered inacceptor stem recognition (8,9). As an extension of this,
agenetic selection has been employed to isolate compensatingmutations in
GlnRS that now aminoacylate (with glutamine) theopal suppressor derived
from tRNA(Gln).The mutations in GlnRSwhich confer altered recognition of
opal suppressor are locatedin two regions. One mutant altered in acceptor
stem recognitionwas previously isolated by a genetic selection scheme using
ambersuppressor tRNAs (8). The second type of mutation, suprisingly,alters
a region that interacts with the inside of the L-shapedtRNA molecule, but
was not implicated by the tRNA(Gln). GlnRSstructure as being important for
specific recognition. Thisregion may provide a structural connection
between the anticodonbinding domain and acceptor stem binding domain of
GlnRS.Analysis of a number of mutations in this region isolated by
acombination of in vitro mutagenesis and genetic selection revealsthe
permutation of residues that modify the communication betweenanticodon and
acceptor stem recognition in the glutamine system.
Keywords: accurate recognition by E.coli glutaminyl-tRNA synthetases,
- Type of paper
: Summary in proceedings
Title: Transfer RNA recognition by Saccharomyces serevisiae
seryl-tRNA synthetase
- Authors:
- Weygand-Đurašević, Ivana (57696)
- Soll, Dieter
- Editors
- Blanquet, S.
Proceedings title: 15th International tRNA workshop
Language: engleski
Place: Cap d'Agde
Year: 1993
Pages: from 312 to 312
Meeting: 15th International tRNA workshop
Held: from 05/30/93 to 06/04/93
Summary: Correct recognition of tRNAs by aminoacyl-tRNA synthetases
is aprerequisite for accurate protein synthesis in the cell. In orderto
elucidate how a eucaryotic class II synthetase specificallyrecognizes its
cognate RNA substrates and discriminates againstnon-cognate tRNAs, we have
focused our investigation onseryl-tRNA synthetases (SerRS) from S.
cerevisiae. Its gene hasbeen cloned and sequenced (1) and can be
functionally expressedboth in yeast and in E.coli. This gives us the
advantage of usinga bacterial host for selected experiments.
The functional complementation of two temperature sensitivemutations in the
E.coli serS gene by S. cerevisiae serS showsthat yeast SerRS recognized and
aminoacylates bacterial serinespecific tRNAs in vivo. The co-expression of
two eucaryotic ambersuppressor tRNA genes, encoding either a human or
aSchizosaccharomyces pombe tRNA(Ser), together with the yeastserS gene in a
suitable E.coli host, resulted in suppression ofE. coli nonsense mutations.
Such a recognition of procaryotic andheterologous eukaryotic serine
specific tRNAs by yeast SerRSsuggests the conservation of at least some
serine identityelements in evolution.
E. coli-like yeast SerRS has been constructed by deletion of theregion
encoding the unusual, positively charged 20 amino acidC-terminal extension
in the eucaryotic protein. Our studies showthat this peptide is dispensable
for the viability of the yeastcell.
- Type of paper
: Summary in proceedings
Title: Changes in gene expression in SV40 transformed human cells
- Authors:
- Rubelj, Ivica (108443)
- Pereira-Smith, O.M.
Proceedings title: The Gerontologist
Language: engleski
Place: Washington, USA
Year: 1992
Pages: from 120 to 120
Meeting: The Gerontological Society of America 45th Annual Scientific Meeting
Held: from 11/18/92 to 11/22/92
Summary: Normal human cells stably express the phenotype of
limiteddivision (senescence) in vitro. Spontaneous imortalization hasnot
been reported for these normal cells. However, theintroduction of DNA tumor
viruses such as SV40 into human cellsresults in immortalization at a low
frequency. It is establishedthat SV40 T ag alone is not sufficient to
immortalize humancells and a second event involving cellular genes must
occur. Toidentity the latter, we studied normal human diploidfibroblasts,
IMR-90, transfected with SV40 in the stages ofprecrisis, crisis and
prostcrisis. Northern blot analysis of cdc2indicated that this gene is
overexpressed 2-3 fold in crisiscells. However, this was not reflected at
the protein level.Fibronectin protein and RNA are at higher levels in cells
incrisis whereas cyclin B protein and RNA are decreased. The SDI1coding
gene coding for an inhibitor of RNA synthesis andoverexpressed in senescent
cells also was overexpressed atcrisis. Flow cytrometric analysis indicates
that cells in crisisaccumulate in the G1 phase of the cell cycle. These
resultssuggest that gene regulation in cells in crisis resembles that
ofsenescent cells in some cases but that T ag disrupts theregulation of
other genes such as cdc2. (Supported by NIH grant2 PO1 AGO7123.)
- Type of paper
: Summary in proceedings
Title: Cloning the gene coding for cytoplasmic tyrosyl-synthetase
from yeast
- Authors:
- Sever, Sanja (179365)
- Editors
- Schwienhorst, A.
- Lindermann, B.J.
Proceedings title: Young perspectives on molecular biotechnology
Language: engleski
Place: Gottingen, Njemačka
Year: 1992
Pages: from 22 to 22
Meeting: Young perspectives on molecular biotehnology First International Meeting
Held: from 02/23/92 to 02/29/92
Summary: Accuracy of translation of the genetic message depends on
atleast two steps: correct aminoacylation of every specific tRNA,catalyzed
by cognate aminoacyl-tRNA synthetase, and correctreading of each codon on
the ribosome.
Yeast tyrosyl-tRNA synthetase (TyrRS) is an alpha2 enzyme ofsubunit
Mr=45000. TyrRS can interact both with cognate andnon-cognate tRNA which is
in discrepancy with her biologicalfunction of specific aminoacylation of
tRNA(Tyr). Complexes ofTyrRS with tRNA were studied by gel retardation
assay. Two kindsof stable complexes were identified. The first complex has
thecomposition tRNA(alpha2) and can be formed both with cognate
andnon-cognate tRNA.
The other complex has the complsition of tRNA(alpha2) and can beformed only
with the cognate tRNA(Tyr). In other to determine theenzyme structure, a S.
cerevisiae genomic expression library wasscreened with antibodies against
TyrRS. In this way positiveclones which contain inserts of different size
(o.2-5 kb) wereobtained. Southern blot show that seven of them cross
hybridized.Based on amino acid sequence homology between known sequence
ofTyrRS from different organisms degenerated pools ofoligonucleotides were
used as primers for amplifyling the targetsequence. The amplifield sequence
belongs to mitohondrial TyrRSand will be used as a probe in Southern blot
in aim to conformhomology between clones obtained with antibodies. DNA
sequencingof positive clones is in progress.
- Type of paper
: Summary in proceedings
Title: Tyrosyl-tRNA Synthetase from yeast
- Authors:
- Sever, Sanja (179365)
- Editors
- Putz, J.
- Dotsch, V.
Proceedings title: Young scientist view of molecular biotechnology
Language: engleski
Place: Strasbourg, Francuska
Year: 1993
Pages: from 25 to 25
Meeting: Young scientist view of molecular biotehnology 2nd International meeting
Held: from 02/28/93 to 03/06/93
Summary: Yeast tyrosyl-tRNA synthetase (TyrRS) is an alpha2 enzyme
ofsubunit Mr=45000. TyrRS can interact both with cognate andnon-cognate
tRNA which is in discrepancy with its biologicalfunction of specific
aminoacylation. tRNA(Tyr) from E.coli is notaminoacylated by yeast TyrRS,
and analysis of a number of mutantsas substrates for yeast TyrRS has led to
identification of C1-G72base pair and discrimination base A73 conserved in
all eucaryoticcytoplasmic tRNA(Tyr), as being important for
recognition.Absolute requirement C1-G72 suggest that the yeast
TyrRScontacts this base pare directly, and if so, this contact islikely to
be in the major-groove side of the base pair as theenzymes belonging to the
class II. However, from it is preferenceof aminoacylation at 2-OH over
3-OH, it would, like E. coli TyrRSseem to belong to class I.
In order to determine the enzyme structure, a S. cerevisiaegenomic
expression library were screened with antibodies againstTyrRS. Between nine
positive clones that were obtained, sevenshare homology. Southern blot show
that a positive clone containg4kb is expressed as fusion protein while
another containing 5kbis expressed from late lambda promoters and internal
translated.In order to sequence this two positive clones they were
clonedinto the M13 vectors and sequenced using synthesizedoligonucleotide
probes. The sequence determined from positivethat contain 4kb contains an
open reading frame for 213C-terminal amino acid but the deduced amino acid
sequence doesnot show strong homology with other TyrRS available
sequences.Since the C-terminal part of the positive that contain 5kb
inorder to determine the N-terminal part of the cloned gene.Sequencing is
in progress.
- Type of paper
: Summary in proceedings
Title: Identification of the gene ror protein kinase from
Saccharomyces cerevisiae
- Authors:
- Sever, Sanja (179365)
- Weygand-Đurašević, Ivana (57696)
- Walter, P.
- Fasiolo, F.
Proceedings title: Godišnji sastanak hrvatskih biokemičara
Language: engleski
Place: Zagreb
Year: 1993
Pages: from 103 to 103
Meeting: Godišnji sastanak hrvatskih biokemičara
Held: from 06/17/93 to 06/18/93
Summary: A 4kb yeast DNA fragment containing a putative TyrRS gene
hasbeen recloned into the M13, mp18 and mp19 vectors and sequencedusing
specially synthesized oligonucleotide primers. An openreding frame of 950
bp was found starting with AUG on theposition 900. Comparison of N-terminal
protein sequence deducedfrom the DNA fragment with protein sequences in the
Swissprotbank showed that a gene coding for a serine/threonine kinase
wascloned. For the initial clasiffication of the novel proteinkinase we
screened the catalytic domain database:genbank/EMBL/PIR. The new enzyme
showed the higest homology withkinases from higher eucaryotes: mouse
ribosomal protein S6kinase, human protein related to PhK-gama, rat
calcium/calmodulindependent protein kinase II, ribosomal protein S6 kinase
fromXenopus oocyte. Since only the N-terminal part of the enzyme
thatposseses kinase catalytic domain was sequenced, addilationsequencing is
necessary for correct clasification of the novelkinase.
- Type of paper
: Summary in proceedings
Title: Interaction of yeast tyrosyl-tRNA synthetase with ATP and
tRNA(Tyr)
- Authors:
- Budiša, N.
- Rubelj, Ivica (108443)
- Katić, M.
- Kućan, Željko (24372)
Proceedings title: Godišnji sastanak hrvatskih biokemičara
Language: engleski
Place: Zagreb
Year: 1993
Pages: from 111 to 111
Meeting: Godišnji sastanak hrvatskih biokemičara
Held: from 06/17/93 to 06/18/93
Summary: Tyrosyl-tRNA synthetase (TyrRS) from Saccharomyces
cerevisiae isan alpha2 enzyme (Mr=2*45000) with "half of site"
activity.However, in addition to the specific cognate
alpha2-tRNA(Tyr)complex, it forms unspecific (alpha2)2-tRNA complexes with
lowestaffinity for TyrRS (Km==,32mM, compared with 8mikroM fortyrosine,
0,12mikroM for tRNA(Tyr) and Ki=0,3mikroM forperiodate-oxidized tRNA(Tyr)
as a competetive inhibitor), wasfound to prevent formation of unspecific
complexes. ATP is alsoan effective inhibitor at high concentrations
(Kss=11mM).Obviously, the second binding site must be involved in
theinhibition. It is likely that in the absence of tRNA the twoTyrRS
subunits can simultaneously catalyze the first reactionstep. Namely, the
k(cat) for pyrophosphate exchange (54 1/s,compared with 2 1/s for the
over-all reaction) is substantiallylowered by the binding of tRNA. Hence,
tRNA binding may reduceaccessibility of the second (or both) binding sites
to the othertwo substrates. On the other hand, ATP binding to the second
sitemay inhibit the over-all reaction either by changing theconformation of
the acting active site, or by direct interferencewith the binding of the
distant (i.e. anticodon) part of the tRNAto the other subunit.
- Type of paper
: Summary in proceedings
Title: Mutants of yeast seryl-tRNA synthetase in the proposed
active site
- Authors:
- Weygand-Đurašević, Ivana (57696)
- Lenhard, Boris
- Landeka, Irena
- Soll, Dieter
Proceedings title: 16th International tRNA workshop
Language: engleski
Place: Madison, Wisconsin, USA
Year: 1995
Pages: from 242 to 242
Meeting: 16th International tRNA workshop
Held: from 05/27/95 to 06/01/95
Summary: Seryl-tRNA synthetase (SerRS) from Saccharomyces cerevisiae
is a homodimeric class II aminoacyl-tRNA synthetase. It shares significant
sequence homology with and has the same oligomeric structure as the SerRS
enzymes from prokaryotic sources. In our search for active site of yeast
SerRS , we have performed saturation in vitro mutagenesis of the portion of
SES1 gene encoding the domain which studies on other class II synthetases
have implicated as being important for the enzyme function. Eleven SerRS
mutants, with amino acid substitutions at various positions between
residues 281 and 297, have been obtained. The contribution of particular
amino acid to the specificity of aminoacylation is being analyzed in vivo,
by complementation of yeast SES1 null-allele strain and in vitro, after
purification of the mutant SerRS enzymes from a yeast overproducing strain.
Keywords: seryl-tRNA synthetase mutants
- Type of paper
: Summary in proceedings
Title: Transfer RNA recognition by class II synthetases
- Authors:
- Weygand-Đurašević, Ivana (57696)
- Editors
- Varljen, Jadranka
Proceedings title: Godišnji sastanak hrvatskih biokemičara
Language: engleski
Place: Opatija
Year: 1994
Pages: from 15 to 15
Meeting: Godišnji sastanak hrvatskih biokemičara
Held: from 10/14/94 to 10/15/94
Summary: Correct recognition of tRNAs by aminoacyl-tRNA synthetases
is a prerequisite for accurate protein synthesis in the cell. This family
of enyzmes can be divided into two classes, which differ in structure of
their ATP binding site and in the mode of interaction with tRNA. In order
to elucidate how an eukaryotic class II synthetase specifically recognizes
its cognate RNA substrates and discriminates against non-cognate tRNAs, we
have focused our investigation on seryl-tRNA synthetase (SerRS) from
Saccharomyces serevisiae. Its gene SES1 has been cloned and sequenced and
can be functionally expressed both in yeast and in Escherichia coli. This
gives us the great advantage of combining genetic, biochemical and
molecular biological approach in our studies. In order to investigate
structure/function relationship in SerRS, a number of mutated forms of the
enzyme has been generated by extensive in vitro mutagenesis of SES1 gene.
Mutations were directed to the putative active site of SerRS, and to its
unusual , positively charged C-terminal extension. In vivo characterization
of the mutants have been achieved by complementation of yeast strain with
disrupted SES1. We have also developed the methods for overproduction and
purification of yeast SerRS and its mutants from E. coli and yeast , what
assures a good source for kinetic analysis in vitro.
Keywords: tRNA, seryl-tRNA synthetase, mutagenesis
- Type of paper
: Summary in proceedings
Title: Mutant forms of seryl-tRNA synthetases
- Authors:
- Landeka, Irena
- Lenhard, Boris
- Weygand-Đurašević, Ivana (57696)
- Editors
- Varljen, Jadranka
Proceedings title: Godišnji sastanak hrvatskih biokemičara
Language: engleski
Place: Opatija
Year: 1994
Pages: from 73 to 73
Meeting: Godišnji sastanak hrvatskih biokemičara
Held: from 10/14/94 to 10/15/95
Summary: The aminoacyl-tRNA synthetases play a crucial role in
protein biosynthesis by specifically charging tRNAs with their cognate
amino acids. This process of precise macromolecular recognition can be
altered by introducing structural changes either into the synthetase or the
tRNA. In our search for the active site of Saccharomyces cerevisiae
seryl-tRNA synthetase (SerRS), we have performed in vitro mutagenesis of
the portion of SES1 gene , which studies on other synthetases have
implicated as being important for the enzyme function. Eleven SerRS
mutants, with amino acid substitutions at various positions between
residues 281 and 297, have been obtained. this domain is highky homologous
between seryl-tRNA synthetases from different organisms. The contribution
of different amino acid to the specificity of aminoacylation has been
analyzed in vivo and in vitro. Yeast SerRS carries a C-terminal extension
of 20 amino acids for wich there is no homology in the prokaryotic
seryl-tRNA synthetases. This stretch is primarily composed of basic
residues. To investigate the possible structural and functional role of
this peptide, truncated enzyme has been generated by expression of of the
deleted SES1 gene, constructed by loop-out mutagenesis with synthetic
oligonucleotide. It retauned low level of aminoacylation activity and its
thermal stability is much below the full-length protein. Our results show
that the C-terminal fragment, although dispensable for the cell viability,
as assayed by in vivo complementation, is important for integrating the
overall structure of the protein.
Keywords: seryl-tRNA synthetase, mutagenesis
- Type of paper
: Summary in proceedings
Title: Human versus bacteria: do bacteria win?
- Authors:
- Kućan, Željko (24372)
Proceedings title: Dani Frane Petriša
Language: hrvatski
Place: Cres, Hrvatska
Year: 1995
Pages: from 27 to 29
Meeting: Dani Frane Petriša
Held: from 07/16/95 to 07/20/95
- Type of paper
: Ph.D.
Title: The activity and specificity of tyrosyl-tRNA synthetase
from yeast of Saccharomyces cerevisiae
Faculty: Prirodoslovno-matematički Zagreb
Date of defense: 02/27/91
Language: hrvatski
Number of pages: 95
Summary: Yeast tyrosyl-tRNA synthetase can interact both with
cognate andnon-cognate tRNAs which is in discrepancy with her
biologicalfunction of specific aminoacylation of tRNA(Tyr).
To investigate the factors responsible for the specificity ofthe enzyme,
retardation gel assay and enzyme kinetics were usedinteraction of TyrRS
with DNA has been, also investigated byelectron microskopy.
It has been found that ATP plays a crucial role in specificity ofthe enzyme
and that 4 negative charges of phosphates arenecessary for such effect. No
influence of tyrosine in theinteraction of TyrRS with tRNA has been
observed.
It has also, been found that a subunit of enzyme can bind bothcognate and
non-cognate tRNAs, however for specific binding oftRNA(Tyr) both subunits
are necessary. This supports ourassumption that tRNA(alpha2) represent
functional complex in theaminoacylation reaction.
TyrRS can also bind double strand DNA and longer chains of asingle strand
DNA, but ATP has no influence in these interaction.The enzyme is bound
within the double strand DNA chain.
The elongated shape of TyrRS, revealed by electron microscopy,offers the
explanation for the behavior of the complexes ingradient gel
electrophoresis.
Keywords: aminoacyl-tRNA synthetase, tRNA
- Type of paper
: M.A.
Title: Cloning the gene for cytoplasmic ryrosil-tRNA synthetase
from yeast Saccharomyces cerevisiae
Faculty: Prirodoslovno-matematički Zagreb
Date of defense: 09/24/93
Language: hrvatski
Number of pages: 83
Summary: The cytoplasmic tyrosyl-tRNA synthetase from yeast is
ahomodimer (alfa2) with subunit molecular weight of 44000. Inother to clone
the gene coding for cytoplasmic tyrosyl-tRNAsynthetase from yeast, S.
cerevisiae genomic expression librarywas screened with antibodies against
TyrRS. In this way ninepositive clones which contain inserts of different
size (0.7-5kb) were obtained. Some of the clones were expressed as
fusionproteins with B-galactosidase, while in others the whole proteinwas
internally translated. Deduced amino acid sequence of a newprotein dose not
show strong homology with other tyrosyl-tRNAsynthetases.
Further analysis of the cloned yeast DNA (933bp), revealedanother open
reading frame, carrying the information forN-terminal part of new protein
kinase. The enzyme shows thehighest homology with kinases from higher
eucaryotes: mouseribosomal protein S6 kinase, human protein related to
PhK-gama,rat calcium/calmodulin dependent protein kinase II,
ribosomalprotein S6 kinase from Xenopus oocyte, mouse, human and rat.
Keywords: aminoacyl-tRNA synthetases, gene cloning, DNA sequencing
- Type of paper
: M.A.
Title: Specific spin labeling of DNA
Faculty: Prirodoslovno-matematički Zagreb
Date of defense: 07/03/91
Language: hrvatski
Number of pages: 72
Summary: The possibility of sequence specific spin labeling of DNA
hasbeen examined in this work. Three different spin probes andseveral DNA
fragments from different sources, but of knownsequences, have been used to
determine these effects. The lengthsof the breakage products of 5-end
labeled DNA treated with spinlabels were compared with the length of DNA
scission productsgenerated by Maxam-Gilbert procedure for DNA sequence
analysis.Spin labeling was performed in the buffer containing 25
mMtriethanolamine HCl pH 7.2 and 1 mM EDTA.
Spin labels I and II were able to modify the DNA. Themodification took
place only at the N-7 guanine atoms.
Fragment BamHI/Sall deriving from plasmid pBR322 wasspecifically spin
labeled. The very limited amounts of spin probeII have selectively
alkylated GGTGG sequence in the fragment.
The influence of spermine and magnesium ions on specific spinlabeling has
also been studied. In their presence, the level ofalkylation was decreased.
The stabilization of DNA structure bymagnesium or spermine, directed the
spin label specifically tothe DNA fragment containing GGTGG sequence.
The frequency of alkylation by spin labels of the other fragmentsshowed
preferential, but not exclusive reaction in the continousruns of guanines
in DNA.
Keywords: DNA labeling, spin probes
- Type of paper
: M.A.
Title: The screening of Saccharomyces cerevisiae genomic DNA
library with antibodies against tyrosyl-tRNA synthetase
Faculty: Prirodoslovno-matematički Zagreb
Date of defense: 07/05/92
Number of pages: 91
Keywords: tyrosyl-tRNA synthetase, genomic library, gene cloning, antibodies
- Type of paper
: Mentorship
Title: The activity and specificity of tyrosyl-tRNA synthetase
from yeast Saccharomyces cerevisiae
Faculty: Prirodoslovno-matematički Zagreb
Date of defense: 02/27/91
Number of pages: 96
Author: Rubelj dr. Ivica
Degree level: Ph.D.
- Type of paper
: Mentorship
Title: The cloning of the gene for tyrosyl-tRNA synthetase from
yeast Saccharomyces cerevisiae
Faculty: Prirodoslovno-matematički Zagreb
Date of defense: 09/24/93
Number of pages: 83
Author: Sever mr. Sanja
Degree level: M.A.
- Type of paper
: Mentorship
Title: Specific spin labeling of DNA
Faculty: Prirodoslovno-matematički Zagreb
Date of defense: 07/03/91
Number of pages: 72
Author: Šušić mr. Slavoljub
Degree level: M.A.
- Type of paper
: Mentorship
Title: The screening of Saccharomyces cerevisiae genomic DNA
library with antibodies against tyrosyl-tRNA synthetase
Faculty: Prirodoslovno-matematički Zagreb
Date of defense: 07/05/92
Number of pages: 91
Author: Pešić mr. Nenad
Degree level: M.A.
- Type of paper
: Invited lecture
Title: Origins and perspectives of molecular biology in Croatia
Institution: Matica Hrvatska
Year: 1994
- Type of paper
: Invited lecture
Title: Molecular interpretation of life
Institution: Matica Hrvatska
Year: 1994
- Type of paper
: Invited lecture
Title: Biology, chemistry or philosophy?
Institution: Academicus
Year: 1995