Enzimologija

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Enzimologija 12. Lekcijas
saturs Praktiska enzimologija. Enzimi
diagnostika. Enzimi terapija. Enzimi
tautsaimnieciba. Molekularas biologijas
(biokimijas)katedra 2009/2010. akad.gads
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Praktiska enzimologija 1.
Diagnostiska enzimologija a) aktivitate
organos (prenatala diagnostika) b) asinis
(klinika nosaka apmeram 30 enzimu aktivitates-
svarigakas AsAT, AlAT, LDH),izoenzimi
c) ELISA 2. Enzimi ka farmakopreparati a)
aizvietotajterapija b) hemostazes
modulacija c) viroterapija d)
onkologija e) pretiekaisuma
terapija 3.Enzimi rupnieciba himozins
(rennins), amilaze, lipazes 4. Praktiskas
enzimologijas perspektivas a) liposomas
b) imobilizacija c) rekombinantu
biotehnologija d) genu terapija
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Enzimi prenatalai iedzimto slimibu diagnostikai
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Prenatala iedzimto slimibu diagnostika visbiežak
tiek lietota glikogenožu, glikolipidožu, glikoprot
eidožu un mukopolisaharidožu diagnostikai
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• Indikatorie enzimi asinis
• Aknu indikatorie enzimi
• Aminotransferazes AsAT, AlAT
• Holinesteraze
• Fruktozes -1,6- difosfat aldolaze
• Histidaze
• Sorbitdehidrogenaze
• Sirds indikatorie enzimi
• Laktatdehidrogenaze
• Aspartataminotransferaze
• Kreatinkinaze
• Pankreasa indikatorie enzimi
• Amilaze
• Lipaze
• Eritrocitu enzimi
• Glikozes-6-fosfatdehidrogenaze
• Prostatas indikatorie enzimi
• Skaba fosfataze
• Kaulaudu indikatorie enzimi

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Dažadi organi ieverojami atškiras ar dažu enzimu
daudzumu tajos
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Alkalai fosfatazei ir daudzas izoformas
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Visatrakais infarkta markieris ir kreatinkinazes
MB izoforma (CK2)
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Laktatdehidrogenazes izoenzimu spektrs dažados
organos
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Indikatorie enzimi urina

• Pankreasa enzimi
• amilaze
• Kaulaudu enzimi
• TRACP - tartrata rezistenta skabas fosfatazes
  izoforma
• Urogenitalas sistemas markierenzimi
• ?-Glutamiltransferaze, ?-Glutamilpeptid
  Aminoskabe
• Laktatdehidrogenaze

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Enzimi bioanalitika
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1. Imunoenzimatiskas reakcijas a)
peroksidaze b) alkala fosfataze
c) glikozooksidaze d) beta-
galaktozidaze e) glukozes-6-fosfat-d
ehidrogenaze f) malatdehidrogenaze 2.
Nukleinskabju kimija a) avidina-
biotina konjugati b) fluorescentie
substrati
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Imunoglobulinu analizes metodes
III Imunoanalizes metodes uz cietas fazes
imunoenzimatiskas metodes,
radioimunometodes. 1. Imunometožu
tipi antivielu saistiba (antibody
capture assay) - piemers antivielu
titra noteikšana ar enzimsaistito
imunosorbenta analizi (Enzyme-linked
Immunosorbent Assays) antigena saistiba
(antigen capture assay)
divu antivielu saistiba (two-antibody
sandwich assys) 2. Popularakie ELISA
varianti a) tieša ELISA b)
netieša c) konkurenta tieša
d) konkurenta netieša e) divpuseja
(sandwich) 3. Izpildišanas tehnika
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Imunoenzimatiskas reakcijas (netiešas
ELISA) princips
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Hemostazi modulejošie enzimi
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Asins recešanas faktori vairuma ir
serina proteinazes
Fak-
Nosaukums,
Aktivators Faktora funkcija
Hemofilija tors sinonimi
(ko
faktors aktive) I Fibrinogens
Trombins
Afibrinogenemija II
Protrombins
Ca2,vit.K, Fibrinogens
Hipoprotrombinemija
faktori
V,X III Audu faktors IV Ca2


II,VII,IX,X V Proakcelerins,
X
II,X Iedzimta
parahemofilija Labilais faktors,


Akcelerator globulins VI Neeksiste,
kludas del aktiveto V faktoru nosauca par VI VII
Prokonvertins
Ca2 , vit.K
K vitamina trukums Seruma
protrombina konversijas
akcelerators, Kotromboplastins VIII
Antihemofilais faktors A, IX,
etc. X
Klasiska hemofilija A
Antihemofilais globulins IX
Antihemofilais faktors B, Ca2 ,
vit.K VIII, X
Hemofilija B, Kristmas faktors ,

K vitamina
trukums Plazmas tromboplastina
komponents X Stujarta-Power
factor Ca2,vit.K,
II Iedzimtais K
vitamina trukums
VIII,IX XI
Plazmas tromboplastins XII
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Hemofilija C XII Hagemana faktors
kollagens, XI
Iedzimts trukums

kallikreins XIII Fibrinu
stabilizejošais faktors II
I Iedzimts
trukums Fibrinoligaze
Fibrina recekli velak škidina plasminogens
(plazmins),kuru aktive audu plazminogena
aktivators (Tissue plasminogen
activator,(alteplase), urokinaze, streptokinaze
etc.


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Faktors XIII, fibrinoligaze ir transferaze
EC 2.3.2.13 Accepted name protein-glutamine
?-glutamyltransferase Reaction protein glutamine
alkylamine protein N5-alkylglutamine
NH3 Other name(s) transglutaminase Factor
XIIIa fibrinoligase fibrin stabilizing factor
glutaminylpeptide ?-glutamyltransferase
polyamine transglutaminase tissue
transglutaminase R-glutaminyl-peptideamine
?-glutamyl transferase Systematic name
protein-glutamineamine ?-glutamyltransferase Com
ments Requires Ca2. The ?-carboxamide groups of
peptide-bound glutamine residues act as acyl
donors, and the 6-amino-groups of protein- and
peptide-bound lysine residues act as acceptors,
to give intra- and inter-molecular
N6-(5-glutamyl)-lysine crosslinks. Formed by
proteolytic cleavage from plasma Factor
XIII References 1. Folk, J.E. and Chung, S.I.
Molecular and catalytic properties of
transglutaminases. Adv. Enzymol. Relat. Areas
Mol. Biol. 38 (1973) 109-191. Medline UI
74169138
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Asins recešanas vispareja shema
Stjuarta-Povera faktors
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Enzimi citu slimibu arstešana

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Preparats Ražotajs
Indikacija Registr.gads Glukocerebro
zidaze Genzyme Goše slimiba
1994 DN-aze (pulmozims) Roche
Cistiska fibroze
1994 Superoksiddismutaze Nippon
Sirds slimibas III F.
1994 Rasburikaze Sanofi
Podagra
2002 alfa-galaktozidaze Genzyme
Fabri slimiba 2001 beta-
galaktozidaze Genzyme Fabri
slimiba 2001 Glukozaminoglikan
- Genzyme Mukopolisaharidoze
2003 iduronidaze Asparaginaze Tripsins Pepsin
s Lipazes Himotripsins Ribonukleaze
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Glikolipidožu terapijai ari megina
lietot trukstošos enzimus
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Enzimi tautsaimnieciba
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Lignins Augstmolekulars zarots polimers no
fenilpropionskabes atliekam (C6-C3)
3 2 1
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Lignin is a complex polymer of phenylpropanoid
subunits. It is an essential component of woody
tissue, to which it imparts structural rigidity.
Lignin is remarkably resistant to degradation by
most microbes nevertheless, a few species of
white-rot fungi are able to catalyse its
oxidation to CO2. Its biodegradation is of great
ecological significance because, next to
cellulose, lignin is the most abundant renewable
polymer on Earth. The first step in lignin
degradation is depolymerization, catalysed by the
lignin peroxidase isozymes (ligninases). These
isozymes are secreted, along with hydrogen
peroxide (H2O2) by the fungus Phanerochaete
chrysosporium Burds. under conditions of nutrient
(nitrogen) limitation. Ligninases are not only
important in lignin biodegradation, but are also
potentially valuable in chemical waste disposal
because of their ability to degrade environmental
pollutants. We have undertaken the cloning of the
ligninase genes to understand further their
regulation and enzymology. We report here the
isolation and characterization of a ligninase
complementary DNA clone with a full-length
insert. The cDNA sequence shows that the sequence
of the mature ligninase is preceded by a
28-residue leader, and the mature protein is
predicted to have a relative molecular mass of
37,000 . Consistent with the classification of
ligninase as a peroxidase certain residues
thought to be essential for peroxidase activity
can be identified and near these residues the
ligninase shows homology with other known
peroxidases. Our cDNA clone has also allowed us
to show that expression of ligninase is regulated
at the messenger RNA level.
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Grutibas enzimu pielietošana un to parvarešana
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Enzimu biotehnologija 1.
Mikrobiala biosinteze 2. Rekombinantu
biotehnologija 3. Virzita mutageneze 4. Enzimu
imobilizacija
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