6_Enzim kul 180310
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Transcript of 6_Enzim kul 180310
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Enzim
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History of enzyme
Energy and life
How enzyme work
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Cells and Energy Chapter 5
~Flow of Energy in Living Things
~The Laws of Thermodyamics
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What is Energy?
Capacity to do work
Forms of energy Potential energy
Kinetic energy
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5.1 The Flow of Energy
in Living Things
Energy
Kinetic and potential
Fig. 5.1
Potential
Energy =storedenergy
Kinetic
Energy =energy of
motion
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5.1 The Flow of Energy
in Living Things
Forms of energy:
Mechanical, sound, light, electric, heat, etc.
Thermodynamics
Study of energy
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5.1 The Flow of Energy
in Living Things
Photosynthetic organisms
SUN
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5.2 The Laws of Thermodynamics
The First Law Energy cannot be created or destroyed
Energy is converted
Heat energy
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Second Law of
Thermodynamics
No energy conversion is ever 100 % efficient
High quality energy is converted to low quality
energy.
Life is a constant battle against the second law.
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The Second Law of Thermodynamics
Disorder increases
Entropy
Fig. 5.3
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Entropy
Measure of degree of disorder in a system
The world of life can resist the flow towardmaximum entropy only because it is
resupplied with energy from the sun
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5.3 Chemical Reactions
Chemical reactions Reactants(substrates)Products
Exergonic reactions
Endergonic reactions
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Energy of Activation
Fig. 5.4
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Catalysts
Fig. 5.4
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Catalysts
speed up the rate of the reaction.
lowers the energy input required for achemical reaction to happen
remains unchanged at the end of thereaction
skool
More free powerpoints at www.worldofteaching.com
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The study history of
enzymes
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The discovery of enzymes as thebiocatalysts (1)
Both enzymology and biochemistry wereevolved from the 19th centuryinvestigation on the nature ofanimaldigestion and fermentation.
Biochemical reactions could not bereproduced in the lab initially and wasthought (e.g., Louis Pasteur) to occur by
the action of a vital force.
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The discovery of enzymes as thebiocatalysts (1)
The idea of catalytic force or contactsubstance promoting fermentation wasintroduced in about 1830s.
Addition of alcohol to an aqueous extractof malt (geminating barley) and salivaprecipitated a flocculent () materialwhich liquefied starch paste and convertedit into sugar, this material was nameddiastase(1833) (lateramylase).
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The discovery of enzymes as thebiocatalysts (2)
Pepsin was discovered as the activeprinciple in the acid extract of gastricmucosa causing the dissolution of
coagulated egg white (1834). Other soluble ferments discovered in the
19th century include trypsin (1857), invertin
(later invertase and sucrase, 1864), papain(vegetable trypsin, 1879), etc.
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The discovery of enzymes as thebiocatalysts (2)
Enzyme (something in yeast) wasfirst coined for such unorganizedferments by Khne in 1876.
Enzymes for alcoholic fermentationwere found to be active in cell freeextracts of yeast (1897, Eduard
Buchner): fermentation is a chemicalprocess, not a vital process.
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The discovery of enzymes as thebiocatalysts (3)
Relationship of initial velocity (V0) andsubstrate concentration (S) wasexamined.
A mathematical description wasestablished for the kinetics of enzymeaction (Michaelis and Menten, 1913).
Th di f th
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The discovery of enzymes as thebiocatalysts (3)
Weak-bonding interactions between the enzymesand their substrates were proposed to distort thesubstrate and catalyze a reaction (Haldane, 1930s).
A GermanA Canadian
A BritishGeneticist
Before it was known that enzymes are proteins!!!
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The Nobel Prize in Chemistry 1907
"for his biochemical
researches and his discoveryof cell-free fermentation"
Eduard Buchner
Germany
Landwirtschaftliche Hochschule
(Agricultural College) Berlin,
Germany
b. 1860, d. 1917.
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Enzyme specificity was revealedby studying sugar conversion
(Emil Fischer, 1890S) Sugars of known structure were synthesized and
used as substrates of enzymes.
The -methylglucoside was found to behydrolyzed by invertin, but not by emulsin,whereas the -methylglucoside was cleaved byemulsin, but not by invertin: the enzyme and the
glucoside was considered to fit (complement)each other like a lock and a key.
Formation of an ES complex was proposed (1894)
E f d b
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Enzymes were found to beproteins
The question of homogeneity of the enzyme preparationsfrustrated the field of enzymology for many decades.
Nitrogen content analysis and various color tests (forproteins) led to contradictory results.
Filterable coenzymes (co-ferments) were discovered inBuchners zymase (Harden and Young, 1906).
Enzymes were thought to be small reactive moleculesadsorbed on inactive colloidal material, including proteins
( as by R. Willsttter in the 1920s). Urease (1926, Sumner) and pepsin (1930, Northrop) were
crystallized and found to be solely made of proteins.
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Urease crystals( X 728)
Sumner, J. B. (1926) Theisolation and
crystallization of theenzyme urease J. Biol.
Chem. 69:435-441.
P i t l
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Pepsin crystals(X90)
Northrop, J. H. (1930)Crystallin pepsin, 1:Isolation and tests ofpurity J. Gen . Physiol.13:739-766.
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The Nobel Prize in Chemistry 1946
for his discovery
that enzymes
can becrystallized"
"for their preparation of enzymes andvirus proteins in a pure form"
James BatchellerSumner John Howard Northrop Wendell MeredithStanley
1/2 of the prize 1/4 of the prize 1/4 of the prize
Cornell UniversityIthaca, NY, USA
Rockefeller Institute forMedical ResearchPrinceton, NJ, USA
Rockefeller Institute forMedical ResearchPrinceton, NJ, USA
1887-1955 1891-1987 1904-1971
Not all enzymes are proteins: Some RNA
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Not all enzymes are proteins: Some RNAmolecules (ribozymes) were found to be
catalytic (Sidney Altman and Thomas Cech,
1981).Ribozymes are found to promote RNA
processing.
Sidney Altman visiting PKU
F ti f b t t
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Formation of an enzyme-substrate(ES) complex was suggested
The activity of invertase in the presence of sucrosesurvives a temperature that completely destroys it if thesucrose is not present (C OSullivan and F. W. Tompson,
1890).
Emil Fishers study on the specifity of invertase (1894).
The rate of fermentation of sucrose in the presence ofyeast seemed to beindependent of the amount of sucrosepresent, but on the amount of the enzyme (A. J. Brown,1902).
The kinetics of enzyme action was originally studied usinginvertase (a hyperbola when V0was plotted against [S]).
The enzyme (E) was thus assumed to form a complex. (ES)with the substrate (S) before the catalysis.
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The kinetics of the enzyme-catalyzed reaction were found to
be rather different from those of atypical chemical reaction
The rate is proportional to the
concentration of the reactantin a typical chemical reaction.
Enzymes however showed
a saturation kinetics:formation of ES complexwas hypothesized (1902).
Th th f lif
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The enzyme theory of life wasformulated
Enzymes are central to every biochemicalprocess (Hofmeister, 1901): life is short andthus has to be catalyzed.
Isolation, purification and physico-chemical
characterization of enzymes would beimportant for understanding the nature of life. Without catalysis, the chemical reactions
needed to sustain life could not occur on a
useful time scale. Self replication and catalysis are believed to be
the two fundamental conditions for life to beevolved. (RNA is thus proposed to be the type
of life molecules first evolved).
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What is enzyme?
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Enzymes
Enzymes are Biological catalysts
Enzymes control chemical reactionsthat take place in the cytoplasm.http://programs.northlandcollege.edu/biology/Biology1111/animations/enz
yme.html
Catalase in an example of an enzymemade by living cells
http://programs.northlandcollege.edu/biology/Biology1111/animations/enzyme.htmlhttp://programs.northlandcollege.edu/biology/Biology1111/animations/enzyme.htmlhttp://programs.northlandcollege.edu/biology/Biology1111/animations/enzyme.htmlhttp://programs.northlandcollege.edu/biology/Biology1111/animations/enzyme.html -
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Susunan enzim
Komponen utama enzim adalah protein Protein yang sifatnya fungsional, bukan
protein struktural
Tidak semua protein bertindak sebagaienzim
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Sifat enzim
Enzim dibentuk dalam protoplasma sel
Enzim beraktifitas di dalam sel tempat
sintesisnya (disebut endoenzim) maupun
di tempat yang lain diluar tempat
sintesisnya (disebut eksoenzim)
Sebagian besar enzim bersifat endoenzim
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How enzyme works?
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Substrate Binding
Unlike inorganic catalysts,enzymes are specific
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Unlike inorganic catalysts,enzymes are specific
succinic dehydrogenase
HOOC-HC=CH-COOH HOOC-CH2-CH2-COOH+2H
fumaric acid succinic acid
NOT a substrate for the enzyme:
1-hydroxy-butenoate: HO-CH=CH-COOH
(simple OH instead of one of the carboxyl's)
Maleic acid
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Dlm system biologi reaksi
kimia selalu memerlukan katalis.
Tanpa katalis sangat lama
shg diperlukan Enzim ygberfungsi sbg biokatalisator
protein yang berfungsi untuk
mempercepat reaksi dengan jalan
menurunkan tenaga aktivasi dantidak mengubah kesetimbangan
reaksi, serta bersifat sangat
spesifik.
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Sifat enzim
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Katalis yg paling
efisien mampu
mempercepat reaksi1020 kali lbh cepat
Enzim bersifat sangat
spesifik, baik jenisreaksi maupun
substratnya ,
Tripsin
Trombin
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Enzim tidak ikut bereaksi dgn substrat atauproduknya (struktur enzim tidak berubah baiksebelum dan sesudah reaksi tetap)
Aktifitas dapat dikontrol sesuai dengankebutuhan organisme itu sendiri
Contoh : enzim yg mengkatalisis reaksipertama pada suatu siklus biosintesis biasanya
di hambat oleh produk akhirnya(feedbackinhibition)
Bagian enzim yang aktif adalah sisi aktif darienzim
bbrp enzim disintesis dlm btk tidak aktif. Danakan diaktifkan oleh kondisi dan waktu yangsesuai (enzim allosterik) . prekursor yg tidakaktif disebutzymogen
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Tiga sifat utama enzim :
Kemampuan katalitiknya
Spesifisitas
Kemampuan untuk diatur (regulasi)
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Bagian-bagian enzim
Beberapa stilah:
Holoenzim
Apoenzim/ apoprotein
Gugus prostetik
Koenzim
Kofaktor
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Bagian-bagian enzim (lanjutan)
Seperti halnya protein lain, enzim memiliki BMantara 12,000 1 juta kd
Beberapa enzim tidak membutuhkan molekulkimiawi lain untuk aktifitasnya, beberapamembutuhkan kofaktor / koenzim
Kofaktor ion-ion inorganik yg dibutuhkanenzim untuk melakukan fungsinya
Koenzim
molekul organik (komplek) yangdibutuhkan enziim untuk melakukan fungsinya
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Protein Enzim protein
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Enzim
Protein psederhana
Protein +Bukan Protein
Protein = apoenzim
EnzimKonjugasi
Bukan protein =
Gugus prostetik
Organik =
Koenzim
Anorganik = kofaktor
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f
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Klasifikasi enzim
Klas Tipe reaksi
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Oksidoreduktase
(nitrat reduktase)
memisahkan dan menambahkan elektron atau
hidrogen
Transferase
(Kinase)
memindahkan gugus senyawa kimia
Hidrolase
(protease, lipase,
amilase)
memutuskan ikatan kimia dengan
penambahan air
Liase
(fumarase)
membentuk ikatan rangkap dengan
melepaskan satu gugus kimia
Isomerase
(epimerase)
mengkatalisir perubahan isomer
Ligase/sintetase
(tiokinase)
menggabungkan dua molekul yang disertai
dengan hidrolisis ATP
Polimerasetiokinase menggabungkan monomer-monomersehin a terbentuk olimer
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Reaksi tanpa enzim:
Lambat
Membutuhkan suhu yang tinggi
Tekanan yang tinggi
Reaksi enzimatis
Enzim memberikan suatu lingkungan yg
spesifik di dalam sisi aktifnya, sehingga reaksisecara energetik dapat lebih mudah terjadi
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Perbedaan antara energi reaktan (fase awal)
dgn energi produk (fase akhir) selisih energi
bebas standar (
G)
Agar reaksi berjalan
spontan, bagaimanakah
nilai G
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Enzim mempercepat reaksi tetapi tidak mengubah
keseimbangan reaksi atau G
Kesetimbangan reaksi antara Reaktan dan produk
mencerminkan perbedaan energi bebas pada fase awal
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Kecepatan reaksi tergantung energi aktifasi G suatu pasokan energi dibutuhkan untuk mengawali suatu
reaksi
Energi aktifasi untuk reaksi yg dikatalis dengan ensim lebihrendah dr reaksi tanpa ensim
Glukosa + 6 O2 6 CO2 + 6 H2O G = -2880 kJ/mol
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Enzim penting untuk menurunkan energi aktifasi untuk
memulai suatu reaksi
Enzim mengikat substrat menciptakan jalan reaksi yg
berbeda yg mempunyai fase transisi lebih rendah dbandingreaksi tanpa enzim
Inti dr reaksi katalisis ikatan yg spesifik pd fase transisi
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Mekanisme kerja enzim
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Fig. 5.5 Enzyme shapes determines its activity
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g y p y
Lysozyme
Changes upon
binding of the
substrate
The substrate is
now bound more
intimately
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Gambar sisi aktif ensim dan asam amino yang
terlibat
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Sisi aktif mempunyai 2 bagian yg penting:
Bagian yang mengenal substrat dan
kemudian mengikatnya
Bagian yang mengkatalisis reaksi, setelah
substrat diikat oleh enzim
Asam amino yang membentuk kedua
bagian tersebut tidak harus berdekatan
dalam urutan secara linear, tetapi dalamkonformasi 3D mereka berdekatan
Teori untuk menjelaskan kerja
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Teori untuk menjelaskan kerja
enzim:
Lock and Key analogyEnzim memiliki struktur sisi spesifik yang cocokdengan substrat.
Mampu menerangkan spesifitas ensim ttp tidak dapat
menerangkan stabilitas fase transisi ensim Induced Fit theory
mempertimbangkan fleksibilitas protein, sehinggapengikatan suatu substrat pada enzim menyebabkansisi aktif mengubah konformasinya sehingga cocok
dgn substratnya. dpt menerangkan fase transisi ESkomplek
Lock and Key
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Lock and Key
http://www.lewport.wnyric.org/jwanamaker/
animations/Enzyme%20activity.html
http://www.lewport.wnyric.org/jwanamaker/animations/Enzyme%20activity.htmlhttp://www.lewport.wnyric.org/jwanamaker/animations/Enzyme%20activity.htmlhttp://www.lewport.wnyric.org/jwanamaker/animations/Enzyme%20activity.htmlhttp://www.lewport.wnyric.org/jwanamaker/animations/Enzyme%20activity.html -
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Enzyme activity
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Enzyme activity
Temperature and pH affect the activityof an enzyme.
E d H
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Enzyme Activity and pH
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Optimum Condition
Enzymes function best or are most activein specific conditions known as optimum
conditions.
Kinetika Reaksi Enzimatis
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Kinetika Reaksi Enzimatis
E + S ES E + PK1 K2
K-1
K1 : kecepatan konstan pembentukan ES komplek
K2 : kecepatan konstan konfersi ES komplek ke PK-1 : kecepatan konstan pemecahan ES komplek ke E
bebas
Enzim sangat efisien dalam mengkatalis suatu reaksi,
steady state(keseimbangan reaksi) segera dapat tercapaiapabila : Kecepatan pembentukan ES komplek sama
dengan kecepatan pemecahannya
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K-1 + K2
= Km konstanta MichaelisK2
Vmax [S]V = Persamaan Michaelis-Menten
Km + [S]
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Lineweaver-Burk double reciprocal plot
Y = m x + b
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penghambatan competitive
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penghambatan competitive
inhibitor bersaing dgnsubstrat untuk terikat pdsisi aktif
Biasanya inhibitorberupa senyawa yg
menyerupai substratnya,& mengikat enzimmembentuk komplek EI
krn terikat scrreversible penghambatan nya
bias, yaitu ketikaditambah substrat makapenghambatanberkurang
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Penghambatan un-competitive
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Penghambatan un-competitive
Terikat pd sisi selain sisiaktif enzim
Berbeda dgn non-competitive inhibitor
ini hanya terikat pd ESkomplek
Sehingga tidak terikat pdenzim bebas
Vmax
berubah, dan Kmjuga berubah
E i ll t ik
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Enzim allosterik
Enzim allosterik mengalami perubahan konformasi sebagai respon terhadap pengikatan modulator efektor
Allosterik enzim biasanya lebih komplek dari nonallosterik enzim, memiliki sub unit lebih dari satu
Memiliki satu atau lebih sisi allosterik / regulator untukmengikat modulator.
Seperti halnya substrat, setiap regulator memiliki sisipengikatan yang berbeda
Untuk enzim homotropik sisi aktif dan sisi regulatorsama
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1. Enzim bersifat koloid, luas permukaan besar, bersifathidrofil
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hidrofil
2. Dapat bereaksi dengan senyawa asam maupun basa,kation maupun anion
3. Enzim sangat peka terhadap faktor-faktor yangmenyebabkan denaturasi protein misalnya suhu, pH dll
4. Enzim dapat dipacu maupun dihambat aktifitasnya
5. Enzim merupakan biokatalisator yang dalam jumlah
sedikit memacu laju reaksi tanpa merubahkeseimbangan reaksi
6. Enzim tidak ikut terlibat dalam reaksi, struktur enzimtetap baik sebelum maupun setelah reaksiberlangsung
7. Enzim bermolekul besar
8. Enzim bersifat khas/spesifik
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specific activity is the amount of product formed by an enzyme
http://en.wikipedia.org/wiki/Enzymehttp://en.wikipedia.org/wiki/Enzyme -
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spec c act ty p y yin a given amount of time under given conditions per milligram ofenzyme.
The rate of a reaction is the concentration of substrate
disappearing (or product produced) per unit time (molL 1s1)
The enzyme activity is the moles converted per unit time (rate reaction volume). Enzyme activity is a measure of quantity ofenzyme present. The SI unit is the katal, 1 katal = 1 mol s-1, butthis is an excessively large unit. A more practical value is 1enzyme unit (EU) = 1 mol min-1 ( = micro, x 10-6).
The specific activity is the moles converted per unit time perunit mass of enzyme (enzyme activity / actual mass of enzymepresent). The SI units are katal kg-1, but more practical units aremol mg-1 min-1. Specific activity is a measure of enzyme
efficiency, usually constant for a pure enzyme. If the specific activity of 100% pure enzyme is known, then an
impure sample will have a lower specific activity, allowing purityto be calculated.
The % purity is 100% (specific activity of enzyme sample /specific activity of pure enzyme). The impure sample has lowerspecific activity because some of the mass is not actually
http://en.wikipedia.org/wiki/Enzymehttp://en.wikipedia.org/wiki/Enzymehttp://en.wikipedia.org/wiki/Enzymehttp://en.wikipedia.org/wiki/Enzyme -
7/30/2019 6_Enzim kul 180310
95/96
-
7/30/2019 6_Enzim kul 180310
96/96