Homology modeling of structure of NH₂-terminal module of mammalian (Bos taurus) tyrosyl-tRNA synthetase
Mammalian tyrosyl-tRNA synthetase (TyrRS) is composed of two structural modules: the NH2-terminal catalytic core and cytokine-like COOH-terminal module. In order to elucidate the structural bases for the N-module functions we have used the computational prediction of its three-dimensional (3D) struc...
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nasplib_isofts_kiev_ua-123456789-1563152025-02-09T22:56:30Z Homology modeling of structure of NH₂-terminal module of mammalian (Bos taurus) tyrosyl-tRNA synthetase Моделювання загомологією структури NH₂-кінцевого модуля тирозил-тРНК синтетази ссавців (Bos taurus) Моделирование по гомологии структуры NH₂-концевого модуля тирозил-тРНК синтетазы млекопетающих (Bos taurus) Odynets, K.A. Bazylevskyi, O.E. Kornelyuk, A.I. Короткі повідомлення Mammalian tyrosyl-tRNA synthetase (TyrRS) is composed of two structural modules: the NH2-terminal catalytic core and cytokine-like COOH-terminal module. In order to elucidate the structural bases for the N-module functions we have used the computational prediction of its three-dimensional (3D) structure by comparative modeling approach. A model of the bovine TyrRS N-module represents the Rossmann nucleotide-binding fold (RF) which is linked to the α-helical domain (αHD). The RF domain forms a single β-sheet containing 5 parallel and one attached antiparallel β-strands surrounded by α-helices. The connective polypeptide, CP1, inserted between β3- and β4-strands of the RF domain is perturbed from the domain core. Comparative analysis of this multiple sequence alignment of known TyrRSs and the obtained model structure reveals the conservative surface elements, which could potentially form the tRNATyr-binding surface. This putative surface includes some exposed amino acid residues of CP1, e. g. essential Lys146 and Lys147 residues, which were identified earlier by site-directed mutagenesis. Тирозил-тРНК синтетаза ссавців (TyrRS) складається з двох структурних модулів: каталітичного NH2-кінцевого кора та цитокінподібного COOH-кінцевого модуля. Для вивчення структурних основ функціонування каталітичного N-модуля Здійснено комп' ютерне моделювання його просторової (3D) структури. Модель структури N-модуля TyrRS являє собою нуклеотидзв'язуючий фолд – згортку Россмана (RF), до якого приєднаний α-спіральний домен (αHD). RF-домен формує β-згортку, яка містить п'ять паралельних та один антипаралельний β-стренди, оточені α-спіралями. З'єднувальний поліпептид, CPі, розміщений між β3- та β4-стрендами RF-домену. Аналіз моделі виявив консервативні елементи, які можуть формувати область зв'язування гомологічної tRNATyr Ця область містить деякі амінокислотні залишки CP1, наприклад, функціонально важливі Lys146 та Lys147, які були ідентифіковані раніиіе методом сайт-спрямованого мутагенезу. Тирозил-тРНК синтетаза млекопитающих (TyrRS) состоит из двух структурных модулей: каталитического NH2-концевого кора и цитокинподобного COOH-концевого модуля. Для изучения структурных основ функционирования каталитического N-модуля проведено компьютерное моделирование его пространственной (3D) структуры. Модель структуры N- модуля TyrRS представляет собой нуклеотидсвязывающий фолд – свертку Россмана (RF), к которому присоединен α-спиральний домен (αHD). RF-домен формирует β-лист, содержащий пять параллельных и один антипараллельный β-стрэнды, окруженные α-спиралями. Соединительный полипептид, CP1, размещен между β3- и β4-cmpендами RF-домена. Анализ модели выявил консервативные элементы, способные формировать область связывания гомологичной tRNATyr . Эта область включает некоторые аминокислотные остатки CP1, например, функционально важные Lys146 и Lys147, которые были идентифицированы ранее методом сайт-направленного мутагенеза 2002 Article Homology modeling of structure of NH2-terminal module of mammalian (Bos taurus) tyrosyl-tRNA synthetase / K.A. Odynets, O.E. Bazylevskyi, A.I. Kornelyuk // Вiopolymers and Cell. — 2002. — Т. 18, № 6. — С. 547-550. — Бібліогр.: 18 назв. — англ. 0233-7657 DOI:http://dx.doi.org/10.7124/bc.000634 https://nasplib.isofts.kiev.ua/handle/123456789/156315 577.152.6:577.332 en Біополімери і клітина application/pdf Інститут молекулярної біології і генетики НАН України |
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Короткі повідомлення Короткі повідомлення |
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Короткі повідомлення Короткі повідомлення Odynets, K.A. Bazylevskyi, O.E. Kornelyuk, A.I. Homology modeling of structure of NH₂-terminal module of mammalian (Bos taurus) tyrosyl-tRNA synthetase Біополімери і клітина |
| description |
Mammalian tyrosyl-tRNA synthetase (TyrRS) is composed of two structural modules: the NH2-terminal catalytic core and cytokine-like COOH-terminal module. In order to elucidate the structural bases for the N-module functions we have used the computational prediction of its three-dimensional (3D) structure by comparative modeling approach. A model of the bovine TyrRS N-module represents the Rossmann nucleotide-binding fold (RF) which is linked to the α-helical domain (αHD). The RF domain forms a single β-sheet containing 5 parallel and one attached antiparallel β-strands surrounded by α-helices. The connective polypeptide, CP1, inserted between β3- and β4-strands of the RF domain is perturbed from the domain core. Comparative analysis of this multiple sequence alignment of known TyrRSs and the obtained model structure reveals the conservative surface elements, which could potentially form the tRNATyr-binding surface. This putative surface includes some exposed amino acid residues of CP1, e. g. essential Lys146 and Lys147 residues, which were identified earlier by site-directed mutagenesis. |
| format |
Article |
| author |
Odynets, K.A. Bazylevskyi, O.E. Kornelyuk, A.I. |
| author_facet |
Odynets, K.A. Bazylevskyi, O.E. Kornelyuk, A.I. |
| author_sort |
Odynets, K.A. |
| title |
Homology modeling of structure of NH₂-terminal module of mammalian (Bos taurus) tyrosyl-tRNA synthetase |
| title_short |
Homology modeling of structure of NH₂-terminal module of mammalian (Bos taurus) tyrosyl-tRNA synthetase |
| title_full |
Homology modeling of structure of NH₂-terminal module of mammalian (Bos taurus) tyrosyl-tRNA synthetase |
| title_fullStr |
Homology modeling of structure of NH₂-terminal module of mammalian (Bos taurus) tyrosyl-tRNA synthetase |
| title_full_unstemmed |
Homology modeling of structure of NH₂-terminal module of mammalian (Bos taurus) tyrosyl-tRNA synthetase |
| title_sort |
homology modeling of structure of nh₂-terminal module of mammalian (bos taurus) tyrosyl-trna synthetase |
| publisher |
Інститут молекулярної біології і генетики НАН України |
| publishDate |
2002 |
| topic_facet |
Короткі повідомлення |
| url |
https://nasplib.isofts.kiev.ua/handle/123456789/156315 |
| citation_txt |
Homology modeling of structure of NH2-terminal module of mammalian (Bos taurus) tyrosyl-tRNA synthetase
/ K.A. Odynets, O.E. Bazylevskyi, A.I. Kornelyuk // Вiopolymers and Cell. — 2002. — Т. 18, № 6. — С.
547-550. — Бібліогр.: 18 назв. — англ. |
| series |
Біополімери і клітина |
| work_keys_str_mv |
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ISSN 0233-7657. Біополімери і клітина. 2002. Т. 18. № 6
Homology modeling of structure of NH2-terminal
module of mammalian (Bos taurus) tyrosyl-tRNA
synthetase
K. A. O d y n e t s , О . E. Bazy levsky i , A· I . K o r n e l y u k
Institute of Molecular Biology and Genetics, National Academy of Science of Ukraine
150 Acad. Zabolothny Street, Kyiv, 03143, Ukraine
E-mail: kornelyuk@imbg.org.ua
Mammalian tyrosyl-tRNA synthetase (TyrRS) is composed of two structural modules: the NH2-terminal
catalytic core and cytokine-like COOH-terminal module. In order to elucidate the structural bases for the
N-module functions we have used the computational prediction of its three-dimensional (3D) structure by
comparative modeling approach, A model of the bovine TyrRS N-module represents the Rossmann
nucleotide-binding fold (RF) which is linked to the α-helical domain (aHD). The RF domain forms a
single β-sheet containing δ parallel and one attached antiparallel β-strands surrounded by a-helices. The
connective polypeptide, CPl, inserted between β3- and β4-strands of the RF domain is perturbed from the
domain core. Comparative analysis of the multiple sequence alignment of known TyrRSs and the obtained
model structure reveals the conservative surface elements, which could potentially form the tRNATyr-
binding surface. This putative surface includes some exposed amino acid residues of CPI, e. g. essential
LysJ46 and Lysl47 residues, which were identified earlier by site-directed mutagenesis.
Introduction. Tyrosyl-tRNA synthetase (TyrRS, ty-
rosine — tRNA ligase, E. C. 6.1.1.1) catalyzes highly
specific attachment of L-tyrosine to cognate tRNATyr
[1 ]. In mammalia (particularly in bovine, Bos taurus)
two functionally active forms of this enzyme have
been discovered. During intracellular proteolysis as
well as after apoptosis-induced secretion, the C-
module may be cleaved from the «minimal» TyrRS
(N-module) within protease-sensitive linking segment
[4—7]. In bovine TyrRS, the N-module is composed
of about 345 amino acid residues (aa) and consists of
a class I-defining Rossmann fold (RF) domain (about
235 aa) and a second anticodon-recognition α-helical
domain (aHD, about 109 aa) [8 ].
As an approach to elucidate the structural bases
of N-module functions we have used the computa-
tional prediction of its three-dimensional (3D) struc-
ture by the comparative modeling approach. 3D
structures of three eubacterial TyrRSs and one tryp-
tophanyl-tRNA synthetase (TrpRS), determined ex-
perimentally [9—12], were used as structural tem-
plates for the homology modeling procedure.
© K. A. O D Y N E T S , Ο. Ε. B AZYLE VS K YI, A. I. KORNELYUK, 2 0 0 2
Materials and Methods. Sequence of BtTyrRS
was deposited to GenBank/GenPept and Swiss-Prot
databases (accession numbers AAC82467, Q29465).
Search for homologous sequences was performed with
iterative PSI-BLAST service (this and other programs
and servers are described in our review [13]). TyrRS
of 15 eukaryotic organisms (Bos taurus, Homo sapi-
ens^ Mus musculus, Fugu rubripes, Drosophila mela-
Itogastery Anopheles gambiae, Caenorhabditis elegans,
Pneumocystis carinii, Saccharomyces cerevisiae, Schi-
zosaccharomyees pombe, Encephalitozoon Cunieuliy
Candida albicans, Plasmodium falciparum, Arabi-
dopsis thaliana and Nicotiana tabacum) and 17
archaebacteria were analyzed. Multiple sequence alig-
nment was carried out with Clustal W and secondary
structure elements were predicted with the PHD and
the multiprediction the NPS@ server. The coordinates
files were obtained from PDB for crystallographic
structures of three eubacterial TyrRSs: Thermus ther-
mophilus (PDB ID codes 1H3E, 1H3F); Bacillus
(Geobacillus) stearothermophilus (2 T S1, 3 T S1,
4TS1); Staphylococcus aureus (1JII, 1JIJ, 1JIK,
IJIL); and tryptophanyl-tRNA synthetase from B.
stearothermophilus (1D2R, 1I6K, 1I6L, 1I6M). Thre-
547
mailto:kornelyuk@imbg.org.ua
O D V N E T S Κ. Α., BAZYLEVSKYI О. Б., KORNELYUK A. I.
ading servers Bioinbgu and FUGUE [14] were used
to search for the similar structures» The complete
N-module model was built using Modeller 6.2 pro-
gram suite [15]. Total 20 initial models from Mo-
deller were built and optimized using «refine-5»
protocol with default parameters for optimization
procedures. Energy calculation of the models has been
performed using Energy option of Modeller.
Secondary structure assignment was done using
DSSP and STRIDE algorithms. Search for structurally
related domains and their superimposition (both in
pair and multiple structure alignment modes) were
carried out using CE and VAST. The SCOP server
1.61 release was used to find other similar structures
from related families and superfamilies. Quality of the
optimized models was verified using Evall23D, ANO-
LEA and ERRAT servers and the best model was
chosen for the subsequent optimization. The models
optimization with a simulated molecular dynamics
(MD) approach was carried out with Gromacs 3.1.4
program [16].
The model structures were placed into a periodic
rectangular box, filled with SPC water layer. The
minimal d is tances between the prote in and
boundaries of the box were 0.7 nm. Energy minimi-
zation was performed on the proteins using a ste-
epest-descent algorithm with 2 fs integration time step
and a tolerance of 0.01 kJ · mof1 · nm""1 during 10 ps.
Position restrained MD was done to distribute water
molecules, and actual MD was simulated during 1 ns.
The temperature was controlled by coupling to an
external bath of 300 K with coupling time constant of
10 fs. The GROMACS force field was used in this
work.
Solvent accessible surface areas of the proteins
were calculated using GetArea 1.1 server. Pockets and
cavities search and analysis was performed using
castP. Distribution of surface conservative residues
was done using ConSurf 2.0. Structure visualization
and analysis were performed with Deep View (Swiss-
PDB Viewer) 3.7b2 and Protein Explorer 1.299.
Results and Discussion. The iterative PSI-BLAST
search for the M1-P344 segment of ^iTyrRS has
found 32 homologous sequences of the N-modules
from other eukaryotic and archaebacterial TyrRSs,
and about 30 sequences of TyrRSs and TrpRSs
homologous to its aHD domain. The N-module sequ-
ences can be divided into five parts according to their
distinct substructures: the N-subdomain (residues
M1-E33); the first half of Rossmann fold (R34-
Kl 19); CPl insertion (G120-L177); the second half
of RF with junked «KMSSS» catalytic loop (K178-
L235); and aHD domain (D236-P344). The highest
local homology between ^ T y r R S and 77TyrRS is in
the N-subdomain and within L-tyrosine binding H3
α-helix and «KMSSS» loop.
We have modeled the £/TyrRS N-module, con-
taining 340 aa residues (L5-P344) with molecular
mass of 38.5 kDa. The 3D structure of the most
similar (69 identical residues for M1-P344 region,
22.5 % identity) T. thermophilus TyrRS (PDB code
1H3E), determined with 2.9 A resolution, was used
as the best template structure. The target/template
pair sequence alignment has been extracted from
multiple sequence alignment and corrected manually
to optimize the positions of insertions. The locali-
zation of insertions/deletions was also deduced from
inspection of the template secondary structure ele-
ments. As the most significant errors in models are
often due to misalignment of target and template
sequences, we performed careful manual editing of
the alignment, as well as the iterative realignment and
model building. There are eleven indels in the
BfTyrRS N-module in comparison with the template,
and their exact locations may vary slightly. There are
only four short insertions within the RF domain of
BtTyrRS (106ESIG, Y129, 148AG and E227) and seven
short insertions in the template 7YTyrRS structure.
Unlike the RF domain, a weak sequence homo-
logy between the aHD domains of eu- and prokaryo-
tic TyrRSs reveals their significant divergence after
the separation of eukaryotes and archaebacteria from
eubacteria. Different threading methods allowed us to
select the B. stearothermophilus TrpRS structure
(PDB code 1D2R, chain A) as an alternative template
for the £/TyrRS aHD modeling because of its higher
structural similarity. For example, sequence-structure
search with Bioinbgu server predicts BsTrpRS as
better template than ^ T y r R S (Z-scores are 39.0 and
4.3 respectively). That sequence-structure alignment
gives only two insertions (258NGVLAFIRHVL and
302EV) within the bovine sequence compared to the
^ T r p R S aHD domain. We have used the BsTrpRS
aHD domain (K192-D297 region) as an alternative
structure template for the aHD structure modeling.
The initial models of the aHD domain were built with
the Modeller program and connected with the RF
domain into complete two-domain N-module structure
by overlapping their common KMSSS-loop segments.
The best from 20 initial models were optimized in
water environment using restricted molecular dyna-
mics simulation techniques with the Gromacs 3.1.4
program.
To test the validity of the initial and optimized
models, a combination of evaluation criteria was used
to discriminate between the correct and incorrect
models. For the models verification such criteria were
used as interatomic clashes, stereochemical properties
548
HOMOLOGY MODELING OF S T R U C T U R E OF NH2-TERMINAL MODULE
(bond lengths, angles and dihedral angles, peptide
bonds planarity, Ca tetrahedral distortion etc.), po-
sition of residues in the Ramachandran plot, root-
mean-square deviation (RMSD) for Ca-Ca atoms of
model/template structure pairs. Using several evalua-
tion servers we analyzed the following structures: 1)
the templates (1H3E and 1D2R); 2) initial and
refined models of the N-module obtained from Mo-
deller; and 3) the optimized best models obtained by
molecular dynamics simulation using the Gromacs
program. ANOLEA server performs energy cal-
culations on a protein chain, evaluating the non-local
environment of each heavy atom in the molecule. The
energy of each pairwise interaction in this non-local
environment is taken from a distance-dependent kno-
wledge-based mean force potential. Server ERRAT
analyzes the statistics of non-bonded interactions
between different atom types, and a single output plot
gives the value of the error function vs. position of a
9-residue sliding window. The error values give
confidence limits that are extremely useful in making
decisions about model reliability. Regions of
candidate protein structures that are mistraced or
misregistered can then be identified by analysis of the
pattern of nonbonded interactions from each window.
The best model obtained after MD simulation
represents a two-domain protein with solvent acces-
sible surface area 16.3 A2 and 41.7 A3 volume
(Figure). The RMSD between the model and template
structures is 1.65 A for 756 backbone atoms.
A number of analyses were carried out to predict
the elements responsible for tRNA binding ability of
N-module. The RF and aHD domains are arranged
into the N-module forming common tRNA binding
surface, which is located on the same side for all
dimeric «minimal» TyrRSs and TrpRSs. The compa-
rative analysis of multiple sequence alignment of
known TyrRSs and the obtained model structure
reveals the conservative surface elements, which could
potentially form the tRNATyr-binding surface. This
putative tRNA-contacting region includes CPl (resi-
dues T121-L177) inserted into RF domain. We have
analyzed the solvent exposed amino acid residues.
The most exposed residues in the CPl of Z?jTyrRS
N-module are: D122, L125, K127, E128, L131, Y134,
R135, S137, S138, T146, Q142, H143, K146, K147,
K154, Q155 and V156.
The secondary structure elements were defined
with the DSSP and STRIDE programs. The RF
domain of the BtTyrRS N-module (residues Ml-
L235) forms a single β-sheet containing 5 parallel and
one attached antiparallel ^-strands arrange as (βΟ)-
β5-β4-βΙ-β2-β3 and surrounded by α-helices. The
/?-sheet adopts additional structural elements. The RF
Accessible surface view of the model structure for N-module of
bovine tyrosyl-tRNA synthetase. The N-module is shown from the
side of its active site cavity, where catalytic KMSSS loop corresponds
to conservative dark-colored amino acid residues. Two domains of
N-module are the Rossmann fold (left) and the α-helical domain
(right). Colors of surface residues correspond to their conservativity
obtained from the multiple sequence alignment analysis
domain includes the additional N-terminal segment of
the first 33 residues containing α-βΟ-β element, which
is characteristic of all known TyrRSs but is not
homologous to TrpRSs and other class I synthetases.
The characteristic CPl insertion (residues G120-
L177) is located between β3- and /34-strands and
perturbs from the core of the RF domain.
A detailed understanding of the protein function
requires the identification of some conserved amino
acid residues at the protein surface, which may be
responsible for the protein function. The ConSurf
server was used to identify such important regions,
based on the phylogenetic relations between homo-
logous proteins from their multiple sequence align-
ment. Conservativity of the exposed amino acid resi-
dues of the eukaryotic-type N-module is represented
on Figure as color-coded surface of the ZOTyrRS
N-module. The majority of exposed (more than
50 %) and strongly conservative residues are loca-
lized in and around the described surface region.
The functional role of lysine residues K146 and
K147 located in CPl of ^ T y r R S has been previously
studied by site-directed mutagenesis [17]. The re-
placement of both residues with Asn and Tyr, respec-
tively, as well as the substitution of K147 alone,
caused the inactivation of mutant TyrRS in the
tRNATyr aminoacylation reaction. In our model both
Lys residues are exposed and may form contacts with
tRNATyr.
Acknowledgements. This work was supported by
NATO Linkage grant HTECG.LG 974684 and NATO
Computer networking supplement No. 976022 to NA-
TO linkage grant.
Note added in proof. When this article was
549
O D Y N E T S Κ. Α. , BAZYLEVSKYI О. Б", KORNELYUK A. I.
submitted, the article of Yang et aL, (2002) [18] was
published where crystallographic structure of a human
«minimal» TyrRS was described with resolution
1.18 A.
K О. Одинець, О. Є. Базилевський, О. І. Корнелюк
Моделювання за гомологією структури ГШ2-кінцевого модуля
тирозил-тРНК синтетази ссавців (Bos taurus)
Резюме
Тирозил-тРНК синтетаза ссавців (TyrRS) складається з двох
структурних модулів: каталітичного NH2-кінцевого кора та
цитокінподібного COOH-кінцевого модуля. Для вивчення стру-
ктурних основ функціонування каталітичного N-модуля Здій-
снено комп' ютерне моделювання його просторової (3D) стру-
ктури. Модель структури N-модуля TyrRS являє собою нукле-
отидзв'язуючий фолд — згортку Россмана (RF)t до якого при-
єднаний α-спіральний домен (aHD). RF-домен формує β-згор-
тку, яка містить п'ять паралельних та один антипаралель-
ний β-стренди, оточені α-спіралями. З'єднувальний поліпеп-
тид, CPі, розміщений між β3- та β4-cmpeндaмu RF-домену.
Аналіз моделі виявив консервативні елементи, які ηожуть
формувати область зв'язування гомологічної tRNA 3^ Ця
область містить деякі амінокислотні залишки CPlt наприк-
лад, функціонально важливі LysI46 та LysJ47, які були іден-
тифіковані раніиіе методом сайт-спрямованого мутагенезу.
К. А. Одынец, А. Е. Базилевский, А. И. Корнелюк
Моделирование по гомологии структуры NH2-концевого модуля
тирозил-тРНК синтетазы млекопитающих (Bos taurus)
Резюме
Тирозил-тРНК синтетаза млекопитающих (TyrRS) состоит
из двух структурных модулей: каталитического NH2-концево-
го кора и цитокинподобного COOH-концевого модуля. Для
изучения структурных основ функционирования каталитиче-
ского N-модуля проведено компьютерное моделирование его
пространственной (3D) структуры. Модель структуры N-
модуля TyrRS представляет собой нуклеотидсвязывающий
фолд — свертку Россмана (RF)t к которому присоединен а-
спиральний домен (aHD). RF-домен формирует β-лист, содер-
жащий пять параллельных и один антипараллельный β-стрэн-
ды, окруженные а-спиралями. Соединительный полипептид,
CP1, размещен между β3- и β4-cmpендами RF-домена. Анализ
модели выявил консервативные элементы, способные форми-
ровать область связывания гомологичной tRNA . Эта об-
ласть включает некоторые аминокислотные остатки CP1,
например, функционально важные Lys146 и Lys147, которые
были идентифицированы ранее методом сайт-направленного
мутагенеза
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УДК 577.152.6:577.332
Надійшла до редакції 17.09.02
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