\Principles of Biochemistry - Lenhingher - 4ta Edicion.
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Transcript of \Principles of Biochemistry - Lenhingher - 4ta Edicion.
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\Principles of Biochemistry - Lenhingher - 4ta Edicion
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Principles of Biochemistry - Lenhingher - 4ta Edicion
Fmoc-aa
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Principles of Biochemistry - Lenhingher - 4ta Edicion
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Principles of Biochemistry - Lenhingher - 4ta Edicion
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Principles of Biochemistry - Lenhingher - 4ta Edicion
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H2N-R1CH-CO2H + H2N-R2CH-CO2H H2N-R1CH-CONH-R2CH-CO2H + H2O 2.1
eq2
[A][B]
O][AB][HK
En esta reacción de equilibrio, dependiendo de los aminoácidos involucrados, el G0 es aproximadamente de 2 kcal/mol; por lo tanto, la formación de una unión peptídica es energéticamente desfavorable y la reacción, en soluciones acuosas, va a estar muy desplazada hacia los aminoácidos libres. La reacción de hidrólisis no catalizada y a temperatura ambiente tiene un t1/2 de 7 años (2×108 s).
OHABBA 21
1
k
k
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2.2 ABAB 10tke
2.3 AB2/AB 2/1100tke
-191 s 103 k
2/112/1 tke
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A partir de
Se puede calcular la constante de equilibrio. A 25 ºC
Como en el equilibrio la velocidad de formación es igual a la de hidrólisis se puede calcular k1 de velocidad de formación como:
k13×10
-9 0.032 sec-1 M-1 9×10-11 s-1M-1
2.4 ln eqKRTG
2.5 M03.0
BA
AB1eq K
2.6 BAAB
1kt
2.7 ABAB
1
k
t
1[A][B]
AB][M03.0
1
1 k
k
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La vida media de un enlace peptidico a pH 7.0 y 250 ºC es aproximadamente 1 minuto
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CAC'NH2
HR1
O
N
CA
R2H
COOH
H
1.521.33
1.45
115.6
121.9
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0.1
HP
PH
0.1
1010
PH7-3-
0.1
1010
PH03-
Protonación del O para dar la forma cargada positiva del enlace peptídico
Para satisfacer el equilibrio:la concentración de la formaprotonada es 10-9 a pH 7 y 10-2 a pH 0
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Deprotonación del N para dar la forma cargada negativamente del enlace peptídico:Como la constante de equilibrio es 10-15, la reacción es insignificante
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Tabla 1. pKa de los grupos ionizables encontrados en las proteínasa
Grupo pKa
-amino 6.8-8.0 -carboxilo 3.5-4.3 -carboxilo (Asp) 3.9-4.0 -carboxilo (Glu) 4.3-4.5 -guanidino (Arg) 12.0 -amino (Lys) 10.4-11.1 imidazolium (His) 6.0-7.0 tiol (Cys) 9.0-9.5 fenol (Tyr) 10.0-10.3 aTomado de Creighton, T. E., Ed. (1993). Proteins: Structure and Molecular Properties. New York, W. H. Freeman and Company.
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pyrrole
pyridine
En los aminoácidos naturales hay dos tipos de nitrógeno formando partede sistemas aromáticos
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R
N+
N-
H
R
NH
N1
2 3
4
5
R
N+
NH
H
RN
+H
NH
R
N
N-
RN
N-
R
N
NH
R
N-
NH+
7.1-
2
10
H
1
H
H3H1a
K
6.5-
2
10
H
1
H
H1H3a
K 14-102
H1a
K
14-102
H3a
K
3-H
1-H
H2+
H-
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Las constantes microscópicas de la figura anterior fueron calculadadas a partir de datos experimentales. Por datos de NMR se sabe que el N1 es el que se protona predominantente. Más precisamente para histidina libre en solución se establece la relación siguiente
Si se mide la constante macroscópica de equilibrio, que no distingue entre las formas microscópicas 1-H y 3-H, se obtiene el siguiente valor
Reorganizando la ecuación anterior
Lo que muestra que la constante macroscópica es la suma de la constantes microscópicas. Por lo tanto
(1) 4H3
H11H/3H
K
(2) 10H
HH1H3K 6.4
2
a 1
(3) 1013
4.6
22
1
H
HH
H
HHK a
(4) 10 4.6H1H3111
aaa KKK
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(5) 10
343 4.6
221
H
HH
H
HHKa
(6) 104 4.6H1H1
111
aaa KKK
(7) 1051 1.7H1
11
aa KK
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lactamasas, mecanismo
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Equilibrio ácido-base
AH A- + H+
Henderson-Hasselbach
][
][log
AH
Aa
pKpH
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1][
][1
][][
][
A
AHAAH
AQ 1
1][][ QA
AH
QA
AH 11
][][
1
1
1-
111
][
][
QQAH
A
QpKapH 1
101 )( )(1011
pKapHQ
QAH
ApKapH
11
1-
][
][10 )(
QpKapH 1
110 )(
De la misma manera
Fracción de carga negativa
)(1011
pKapHQ
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j
jji
ii QnQnQ
)( pHfQ
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ONH
SCH3
NHR1R2
N
BrH
+
ONH
S+
CH3
NHR1
N
R2
ONH
NR1R2
CH3 SCN
BrH
H2O
NH R2
ONH
R1OH
NH2R2 (péptido con extremo amino libre)
O
NH
R1 O
peptidil homoserinalactona
OH
NH
R1 O
OH
H2O
peptidil homoserina
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1. A partir de los pKa tabulados en los textos calcule la carga neta de los siguientes polipeptidos a pH=7 y a pH=2.
a) ACDALDSRQTGH
b) QNRSKHLLLKDG
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1. A partir de los pKa tabulados en los textos calcule la carga neta de los siguientes polipeptidos a pH=7 y a pH=2.
a) ACDALDSRQTGH
b) QNRSKHLLLKDG
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2. En el diseño de un proceso cromatográfico de intercambio iónico se busca definir la mejor condición para separar dos péptidos. Se dispone de dos alternativas: una resina sulfónica equilibrada a pH 8.0 y una de amino cuaternario equilibrada a pH 5.0. Para eluir se usará un gradiente de fuerza iónica. La estructura de los dos productos a separar es la siguiente:
a) EERAVY
b) HNSTFHWGD
Indique si la separación tiene posibilidades de éxito y cuál de los sistemas cromátográficos usaría.