Download - Cofatores inorgânicos
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Cofatores inorgânicos
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Cofatores orgânicos ou metalo-orgânicos = coenzimas
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Quimotripsina
Sítio ativo
Substrato “modelo”
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Caminho energético de uma reação química não-catalizada
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Caminho energético de uma reação química catalizada por uma enzima
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G = Go + RTln([produtos]/[reagentes])
Reações são espontâneas se G < 0
G é uma função de:
i) Concentração relativa de reagentes e produtosii) Temperaturaiii) Tendência intrínseca da reação ocorrer, expressa explicitamentepelo valor de Keq ou Go
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G = Go + RTln([produtos]/[reagentes])
No equilíbrio: i) G = 0 ii) [produtos]/[reagentes] = Keq
Logo, no equilíbrio:
0 = Go + RTlnKeq
ou
Go = - RTlnKeq
Keq = exp (-Go/RT)
Relação entre Go e Keq
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Go = - RTlnKeq
Keq = exp (-Go/RT)
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Mecanísmos de catálise: vide Voet capitulo 11, parte 3 (seção 11-3)
1) Catálise ácido-base2) Catálise covalente3) Catálise por íons metálicos4) Catálise eletrostático5) Efeitos de proximidade e orientação6) Ligação preferencial do complexo de estado de transição
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Complementaridade entre o sítio ativo e o substrato
Diidrofolato redutase e seus substratos tetraidrofolato e NADP+
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Energia livre do estado de transição é necessária para torçer obastão no caminho até sua quebra
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Energia livre de ligação (GM) do bastão com a enzima neste exemplo estabiliza a forma não torcida do bastão.
Logo a energia livre do estado de transição necessária para torçer o bastão aumenta
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Energia livre de ligação (GM) neste exemplo estabiliza a forma torcida do bastão.
Logo a energia livre do estado de transição necessária para torçer o bastão diminui
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Enzimas utilizam a energia de ligação para diminuir a energia livre do estado de transição.
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Mudanças conformacionais associadas com ligação do substrato
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hexoquinase
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Hexoquinase + D-glicose
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Catálise por redução de entropia
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Catálise porÁcidos e Bases – Uma tema Central paraCatáliseEnzimática
Ácido específico = H3O+
Base específica = OH-
Ácido geral = HABase geral: B:
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Estabilização do estado de transiçãoe catálise por íons metálicos
Catálise básicogeral (remoção de H+)
Catálise ácidogeral (eliminação de OH-)
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Alosteria ou Regulação alostérica
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Aspartato transcarbamoilase
- moduladora + moduladora (ie CTP)
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Autoregulação de viasmetabólicas
Inibição “feedback”
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Hiperbólica: Lembra de curva de ligação de O2 para mioglobina
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Lembra de : Y = [L]/(Kdiss + [L])
Velocidadenaquela [S]
Velocidade máximaConcentraçãodo substrato
Kdiss aparente do Complexo enzima-substrato
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E + S ES E + P
Ks = [E][S]/[ES]= k-1/k1kcat
Velocidade de reação = d[P]/dt = kcat[ES]Podemos assumir que d[ES]/dt = 0 (assunção de estado
estacionário)Logo: taxa de formação de ES = taxa de sua destruição
k1[E][S] = (k-1 + kcat)[ES] k1{[ETOT]-[ES]}[S] = (k-1 + kcat)[ES]
k1[ETOT] [S] - k1 [ES][S] = (k-1 + kcat)[ES] k1[ETOT] [S] = k1 [ES][S] + (k-1 + kcat)[ES] k1[ETOT] [S] = [ES]{k1[S] + (k-1 + kcat)} [ES]= k1[ETOT] [S]/{k1[S] + (k-1 + kcat)} [ES]= [ETOT] [S]/{[S] + (k-1 + kcat)/ k1} [ES]= [ETOT] [S]/{[S] + KM} onde KM = (k-1 + kcat)/ k1
Logo: velocidade = kcat[ETOT][S]/(KM + [S])
k1
k-1
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E + S ES E + P
Ks = [E][S]/[ES]= k-1/k1kcat
velocidade = kcat[ETOT] [S]/(KM + [S])
Vo = kcat[ETOT][S]/(KM + [S])
Vo = Vmax[S]/(KM + [S]) onde Vmax = kcat[ETOT]
Notar que KM = (k-1 + kcat)/ k1
Logo, se k-1 >> kcat KM = k-1/ k1 = Ks = [E][S]/[ES] =Constante de dissociação
do complexo ES (ezima-substrato)
k1
k-1
Condição de equilíbrio rápido Entre E, S e ES.
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Quando[S] >>KM
Quando[S] << KM
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Lineweaver-Burke
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Vo = kcat[ETOT][S]/(KM + [S])
Quando [S] << KM : Vo = kcat[ETOT][S]/KM = (kcat/KM) x [ETOT][S]
Logo, kcat/KM é um aparente constante cinético bimolecular da reação E + S E + P
E tem um limite máximo de 108 – 109 M-1s-1
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= 1 + [I]/KI
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’ = 1 + [I]/K’I
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= 1 + [I]/KI
’ = 1 + [I]/K’I
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Inibição Competitiva
Vo = Vmax[S] KM + [S]
= 1 + [I]/KI
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Inibição não-competitiva(“uncompetitive”)
Vo = Vmax[S] KM + ’[S]
’ = 1 + [I]/K’I
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Inibição Mista
Vo = Vmax[S] KM + ’[S]
= 1 + [I]/KI
’ = 1 + [I]/K’I
(Quando = ’,
“non-competitive”
Intercepto no eixo X)
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Inibidora suicida
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EH+ E + H+ativa inativa
pKa = 3
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EH22+ EH+ + H+ E + 2H+
inativa pKa ativa pKa inativa = 6,5 = 9,2
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Hidrólise de amidas catalizada por quimotripsina
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quimotripsina
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quimotripsina
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qumiotripsina
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Sítio ativo de quimotripsina
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Cooperatividade e alosteria
Exemplo: enzima homotrópica (substrato é um modulador positivo ou ativador)
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Exemplo: enzima alostérica
(moduladores positivos ou negativos mudam KM)
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Exemplo: enzima alostérica
(moduladores positivos ou negativos mudam Vmax)
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REGULAÇÃO POR MODIFICAÇÃO COVALENTE
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