Projects in need of doing

Rui Alves

Metabolic Reconstruction

Rui Alves

To do

• Literature analysis– Semi done– Needs to implement

• FREQUENCY ANALYSIS• CO-OCCURENCE ANALYSIS• A GOOD DATA BASE STRUCTURE TO KEEP

TRACK OF PREVIOUS SEARCHES AND USE THEM

• A GOOD VISUALIZATION TOOL

You would learn

• Literature analysis– PHP– APACHE– MAYBE JAVA AND C

To Do

• Phylogenetic analysis– 60% done– Needs to improve the database structure– Needs to implement a web front to connect

the database to a browser

You would learn

• Literature analysis– PHP– APACHE– HTML– MYSQL

To Do

• MODELLING & DOCKING– IMPLEMENT A METASEARCH TOOLS TO

USE EXISTING MODELLING SERVERS– SAME FOR DOCKING– IMPLEMENT IN HOUSE WAY OF RUNNING

ROBETTA – WEB INTERFACE

You would learn

• Literature analysis– PHP– APACHE– HTML– C

To Do

• INTEGRATIVE ARCHITECTURE– DESIGN ARCHITECTURE TO INTEGRATE

ALL DATA

You would learn

• Literature analysis– PHP– APACHE– HTML– C

To Do

• BIOLOGICAL APPLICATION, ALLWAYS

REWARDS

• CO-AUTORSHIP IN PAPERS DESCRIBING THE WORK

• THERE MAY BE SOME MONEY FOR ONE PART TIME JOB FOR A FEW MONTHS, DEPENDING ON HOW THE MINISTRY ALLOWS THE ADMINISTRATION OF THE PROJECTS

Design Principles in Signal Transduction

Rui Alves

To Do

• Compare MAPK pathways to TCS

• Compare different types of TCS

• Compare different types of MAPKS

TCS MAPK

You would learn

• MATHEMATICA

• MATHEMATICAL MODELLING

REWARDS

• CO-AUTORSHIP IN PAPERS DESCRIBING THE WORK

Modeling the life cycle of the lac operon

Rui Alves

To Do

• Start working with stochastica approximations to analyze demand theory not only in gene expression but also in protein modules

You would learn

• MATHEMATICA

• MATHEMATICAL MODELLING

• Mathematically Controlled Comparisons

• Theoretical ecology

REWARDS

• CO-AUTORSHIP IN PAPERS DESCRIBING THE WORK

• Possibility of going abroad if funding can be secured

The lac operon

lacZ (3510) lacY (780) lacA (825)POlacI (1040)

-Gal Permease AcetylaseInhibitor

lacI

CAP

RNAPol

Maintenance of the lac operon as a function of life style

Can one understand, quantitatively, which are the boundaries for selection of the lac operon so that bacteria do not lose this operon through random drift?

The lac operon

lacZ (3510) lacY (780) lacA (825)POlacI (1040)

-Gal Permease AcetylaseInhibitor

lacI

Mike has done the analysis, with the simplifying assumptions that no glucose was ever in the medium (i.e. CAP is always on) and considering only the regulator mutants and building and analyzing a model of bacterial growth

Determining the mutant to wild type ratios as a function of time

, ,

, ,

w ww w

P P w w P P P

m m w w m m m

X X

X X X

X X X

2

, ,

2

, ,

/ /( ) / ( / ) /

/ /( ) / ( / ) /

P w w PP P w

w

P P ww P P w

m w w mm m w

w

m m ww m m w

dX dt X dX dt Xd R dt d X X dt

X

R

dX dt X dX dt Xd R dt d X X dt

X

R

, ,,

, ,

, ,,

, ,

0

0

P w P wP P P ww P

ww P P ww P P

m w m wm m m ww m

ww m m ww m m

R t Exp t R

R t Exp t R

Bacteria Cycle between environments with high (H) and low

(L) Lac

,,

,

,

,

1

, ,

,

,

,

1

HP w H H

P i P P wwH Hww P P

LP w

L Lww P P

P i

L L LP P ww P w

L Lww P P

HP w

H Hww P P

R C Exp CD

R C

Exp C D

Parameter values are a function of the environment and of the

genotype

Hwg 1(1 )H H H H

ww w w P w m

L Hww ww

g m m

0.0125

Hw P P

Hw m m m

m

m

9 1 16 10

10

30

60

P

m

m

mutations res generation

res

res

res

Selection boundaries for the lac operon

0.0001 0.001 0.01 0.1 1

LogD1

100

10000

1. 106

1. 108

1. 1010

LogCPromoter Ratio= 0.001

Regulator Ratio= 0.001

Selection for inhibitor and promoter elements

The lac operonlacZ (3510) lacY (780) lacA (825)POlacI (1040)

-Gal Permease AcetylaseInhibitor

lacI

CAPRNAPol

, ,

, ,

w w w

wY Y w Y Y Y

wZ Z w Z Z Z

X X

X X X

X X X

, ,

, ,

, ,

, ,

wI I w I I I

wP P w P P P

wP P w P P P

wCAP CAP w CAP CAP CAP

X X X

X X X

X X X

X X X

Xw

XZ-

XY-

XI-

XCAP-

XP-

XP+

The lac operon: Selecting an up-promoter

lacZ (3510) lacY (780) lacA (825)POlacI (1040)

-Gal Permease AcetylaseInhibitor

lacI

CAPRNAPol

Xw

XZ-

XY-

XI-

XCAP-

XP-

XP+

, ,wP P w P P PX X X

Gain of function promoter mutant

0.00001 0.0001 0.001 0.01 0.1LogD1

100

10000

1. 106

1. 108

1. 1010

LogC Stronger promoter selected

The lac operon:Selecting a down-promoter

lacZ (3510) lacY (780) lacA (825)POlacI (1040)

-Gal Permease AcetylaseInhibitor

lacI

CAPRNAPol

Xw

XZ-

XY-

XI-

XCAP-

XP-

XP+

, ,wP P w P P PX X X

Loss of function promoter mutant

0.00001 0.0001 0.001 0.01 0.1LogD1

100

10000

1. 106

1. 108

1. 1010

LogCStronger promoter selected

Weaker promoter selected

Maintenance of current promoter

The lac operon: Selecting for less efficient lacZ

lacZ (3510) lacY (780) lacA (825)POlacI (1040)

-Gal Permease AcetylaseInhibitor

lacI

CAPRNAPol

Xw

XZ-

XY-

XI-

XCAP-

XP-

XP+

, ,wZ Z w Z Z ZX X X

Loss of function -Gal

0.00001 0.0001 0.001 0.01 0.1LogD1

100

10000

1. 106

1. 108

1. 1010

LogC

Weaker galactosidase selected

Maintenance of current -Gal plus promoter

lacZ (3510) lacY (780) lacA (825)POlacI (1040)

-Gal Permease AcetylaseInhibitor

lacI

CAPRNAPol

Xw

XZ-

XY-

XI-

XCAP-

XP-

XP+

, ,wY Y w Y Y YX X X

The lac operon: Selecting for less efficient lacY

Loss of permease function mutant

0.00001 0.0001 0.001 0.01 0.1LogD1

100

10000

1. 106

1. 108

1. 1010

LogC

Weaker permease selected

Maintenance of current permease plus -Gal plus

promoter

lacZ (3510) lacY (780) lacA (825)POlacI (1040)

-Gal Permease AcetylaseInhibitor

lacI

CAPRNAPol

Xw

XZ-

XY-

XI-

XCAP-

XP-

XP+

, ,wI I w I I IX X X

The lac operon: Selecting for less efficient lacI

Loss of function inhibitor

0.00001 0.0001 0.001 0.01 0.1LogD1

100

10000

1. 106

1. 108

1. 1010

LogC Maintenance of current repressor protein plus

permease plus -Gal plus promoter

Weaker repressor selected

lacZ (3510) lacY (780) lacA (825)POlacI (1040)

-Gal Permease AcetylaseInhibitor

lacI

CAPRNAPol

Xw

XZ-

XY-

XI-

XCAP-

XP-

XP+

, ,wCAP CAP w CAP CAP CAPX X X

The lac operon: Selecting for less efficient CAP

Loss of function activator

0.001 0.01 0.1 1LogD1

100

10000

1. 106

1. 108

1. 1010

LogC

Demand for activator OR absence of Glucose

If D~<2x10-4, activator is lost for this particular circuit

Connecting Lac operon demand regulated by Glu and Lac

Demand for LacI

Demand for CAP

Connecting Lac operon demand regulated by Glu and Lac

Demand for LacI

Demand for CAP

Connecting Lac operon demand regulated by Glu and Lac

Demand for LacI

Demand for CAP

Connecting Lac operon demand regulated by Glu and Lac

Demand for LacI

Demand for CAP

Connecting Lac operon demand regulated by Glu and Lac

Demand for LacI

Demand for CAP

Connecting Lac operon demand regulated by Glu and Lac

Demand for LacI

Demand for CAP

Connecting Glu and Lac demand

6

1

1ww w ii

g m

ww

Y Y

Xw

XZ-

XY-

XI-

XCAP-

XP-

XP+

Z Z

P P

P P

I I

CAP CAP

,P w

,P w

,CAP w

,Z w

,Y w

,I w

., .w wg m

6

.,. .1.

1 iii

g m

Bacteria Cycle between environments with high and low Lac and high and low

Glu

,,

,

,

,

, 1 2 3 ,

,

,

,

...

1

HP w H H

P i P P wwH Hww P P

LP w

L Lww P P

L L LP P ww P w

L Lww P P

HP w

H Hww P P

R C Exp CD

Exp C D D D

4 fractions of the cycle

The words:

Demand for LacI

Demand for CAP

Highest extend of selection for this type of demand cycle

~99.2%0.8%

~0.005%~0.07%

HL,HH,HL,LL

-4 -2 0 2 4

3

3.2

3.4

3.6

3.8

LogC

LogExtent of selection

HL,HH,HL,LL

0.992 0.993 0.994 0.995 0.996 0.997

3

3.2

3.4

3.6

3.8

LogFraction of low demand

LogExtent of selection

HL,HH,HL,LL

-4 -2 0 2 4

0.992

0.993

0.994

0.995

0.996

0.997

LogC

LogExtent of selection

HL,HH,HL,LL

-4 -2 0 2 4

0.002

0.004

0.006

0.008

LogC

t1

HL,HH,HL,LL

-4 -2 0 2 4

0

0.00001

0.00002

0.00003

0.00004

0.00005

t2

HL,HH,HL,LL

-4 -2 0 2 4

0

0.001

0.002

0.003

0.004

0.005

0.006

0.007

t3

HL,HH,LH,LL

-4 -2 0 2 4

2.8

3

3.2

3.4

3.6

3.8

LogC

LogExtent of selection

0.976 0.978 0.98 0.982 0.984 0.986

2.8

3

3.2

3.4

3.6

3.8

LogFraction of low demand

-4 -2 0 2 4

0.976

0.978

0.98

0.982

0.984

0.986

Low Demand

-4 -2 0 2 4

0.005

0.01

0.015

0.02

LogC

t1

HL,HH,LH,LL

-4 -2 0 2 4

-1

-0.5

0

0.5

1

LogC

t2

-4 -2 0 2 4

0

0.002

0.004

0.006

0.008

0.01

t3

-2.4 -2.2 -2 -1.8 -1.6

2.8

3

3.2

3.4

3.6

3.8

Logt1

LogExtend of Selection

HL,LL,LH,LL

-4 -2 0 2 4

2.8

3

3.2

3.4

3.6

3.8

LogC

LogExtent of selection

-2.4 -2.2 -2 -1.8 -1.6

2.8

3

3.2

3.4

3.6

3.8

Logt1

LogExtend of Selection

0 0.05 0.1 0.15 0.2 0.25 0.3 0.35

2.8

3

3.2

3.4

3.6

3.8

t2

0.65 0.7 0.75 0.8 0.85 0.9 0.95

2.8

3

3.2

3.4

3.6

3.8

LogFraction of low demand

LogExtent of selection

HL,LL,LH,LL

-4 -2 0 2 4

0.65

0.7

0.75

0.8

0.85

0.9

0.95

LogC

Low Demand

-4 -2 0 2 4

0.005

0.01

0.015

0.02

LogC

t1

-4 -2 0 2 4

0

0.05

0.1

0.15

0.2

0.25

0.3

0.35

t2

-4 -2 0 2 4

0

0.002

0.004

0.006

0.008

0.01

t3

HH,HL,LL,LH

-4 -2 0 2 4

1.2

1.4

1.6

1.8

2

LogExtent of selection

0.38 0.382 0.384 0.386 0.388 0.39

1.2

1.4

1.6

1.8

2

-4 -2 0 2 4

0

0.2

0.4

0.6

0.8

1

1.2

Low Demand

HH,HL,LL,LH

-4 -2 0 2 4

0.38

0.382

0.384

0.386

0.388

0.39

t3

-4 -2 0 2 4

0.01

0.012

0.014

0.016

0.018

0.02

t2

Sensitivity Analysis

-4 -2 0 2 4

2

2.5

3

3.5

4

LogC

LogExtend of Selectionmiu 5

-4 -2 0 2 4

2

2.5

3

3.5

4

LogC

LogExtend of Selectionmiu 4

-4 -2 0 2 4

2

2.5

3

3.5

4

LogC

LogExtend of Selectionmiu 3

-4 -2 0 2 4

2

2.5

3

3.5

4

LogC

LogExtend of Selectionmiu 2

Sensitivity Analysis

-4 -2 0 2 4

2

2.5

3

3.5

4

LogC

LogExtend of Selectionmiu 1

-4 -2 0 2 4

1.8

2

2.2

2.4

2.6

2.8

3

3.2

LogExtend of Selectionmiu 0

Lambda-cro

-5

0

5-8

-6

-4

-2

0

11.000251.0005

1.000751.001

-5

0

5

-5

0

5-8

-6

-4

-2

0

0.999920.999940.999960.99998

1

-5

0

5

-5

0

5-8

-6

-4

-2

0

11.000251.0005

1.000751.001

-5

0

5

-5-2.5

02.5

5 -10

-8

-6

-4

-2

0

0.166660.1666650.16667

0.166675

-5-2.5

02.5

5

The lac operon

High Lactose Low Lactose

High Gluco

se

Low Gluco

se

High Gluco

se

Low Gluco

se