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Saturday, December 13, 2008

Web Source: Biochemical Pathways

BIOCHEMICAL PATHWAYS


Biochemical processes may be conveniently viewed as an ASSEMBLY LINE of SEQUENTIAL CHEMICAL REACTIONS (examine three or more different pathways at this site), each controlled by a unique enzyme. The biochemists have given the various series of enzyme-directed reactions names, like the TCA CYCLE, GLYCOLYSIS etc. (click the pathways to view the animated steps using the helper app Chime). for ease of discussion. To see animations of how specific chemical reactions look click here then click on "Glycolysis" followed by "Glycolysis--Step by Step". However, remember that everything in a cell is INTIMATELY


INTERCONNECTED and every individual reaction must be thought of as affecting every other reaction. One proof that all life is related, is that a core of biochemical pathways run throughout life. The sugar glucose, for example, is used as a food by almost every organism on the planet and the enzymatic pathways for utilizing glucose are extremely similar. The various enzymes involved may differ in physical characteristics, but the chemical steps involved in glucose breakdown are the same. "Do you think that this continuity of life does or does not support the theory of evolution?" Click here to see a metabolic map like that on the wall in the lab. Click on the Carbohydrate Metabolism label and view that portion of the metabolic map.


Although most of the core biochemical pathways are known, some specialized pathways are still unknown: such as those involved in the synthesis of some antibiotics. The major research issue today is how enzyme activity and synthesis are REGULATED. For example, every human cell with a nucleus contains the ENTIRE COMPLEMENT OF HUMAN GENES, yet only a small portion of these genes are active at any time in any given cell; in the eye only genes involved for vision are active, in the liver, only those required for liver cell functioning are active etc. The development of a human from a single egg is a poorly understood process. How specific sub-sets of genes are turned on and off in time and space present an exciting challenge for scientists.


Click here or here for excellent presentations of glycolysis. Click here to have a choice of pathways you want to see. Click here for a great site on pathways and for a colored map showing the interconnectivity of the metabolic pathways.


Click here to view the complete biochemical chart. Type in "glucose 6 phosphate" and hit ENTER. Use the arrows to move around the chart.

SourceDr. R. E. Hurlbert, WSU.EDU



BIOCHEMICAL PATHWAY DIAGRAMS


Stages of Catabolism

Calvin Cycle Flow Diagram

Glycolysis

Glycolysis Enzyme Mechanisms

Glyoxylate Cycle

Aerobic Glucose Metabolism

Fatty Acid Oxidation Reactions

Glycolysis/Gluconeogenesis Pathways

Fatty Acid Synthase Complex

Gluconeogenesis in the Liver

Fatty Acid Biosynthesis Reactions

Pentose Phosphate Pathway

Pyruvate-Malate Cycle

Glycogen Cascade Control

Fatty Acid Regulation

Pyruvate DH Complex Reactions

Kreb's Urea Cycle

Kreb's TCA Cycle

Basic Folate Metabolism Pathways

Kreb's TCA Cycle Intermediate Metabolism

Purine Biosynthesis

Malate-Aspartate Shuttle

Purine Degradation

(Sourece: Richard A. Paselk, Humboldt State University)


Other Resources of Biochemical Pathway


Metabolic pathway - Wikipedia, the free encyclopedia

In biochemistry, a metabolic pathway is a series of chemical reactions occurring within a cell. In each pathway, a principal chemical is modified by ...
en.wikipedia.org/wiki/Metabolic_pathway


Image results for biochemical pathways


ExPASy - Biochemical Pathways

and Pathway Map

Digitized version of wall charts courtesy Boehringer Mannheim et al, divided into Metabolic Pathways and Cellular and Molecular Processes, maintained by the ...
www.expasy.ch/cgi-bin/show_thumbnails.pl


Pathway Tools Query Page

Each dataset contains classification hierarchies for pathways, for reactions ( the enzyme nomenclature system), for compounds, and for genes. ...
pathway.yeastgenome.org/biocyc/


Pathways

Pathways is provided an an ancillary to General, Organic and Biochemistry and Concepts of Biochemistry. Please send any comments or suggestions to Jim Hardy ...
ull.chemistry.uakron.edu/Pathways/index.html


Yeast Biochemical Pathways

Yeast Biochemical Pathways are created using the Pathway tools software developed by Peter Karp and his colleagues at SRI International. ...
www.yeastgenome.org/biocyc/

Friday, December 12, 2008

Biochemical Protocol 1. Basic Lab Technology

Alternate method for purifying DNA from agarose gels

Isolation of RNA from mammalian cells using RNAZOL (Teltest)

Preparation of sequencing gels

Preparing proteins/antibodies/DNA/RNA for microinjection

RT-PCR Protocol

Safety Procedures
Preparation of Solutions
Disposal of Buffers and Chemicals
Equipment

Southern blotting

Transfection of mammalian cells using Lipofectamine (LTI)

Transformation of E. coli by electroporation

Wizard PCR preps DNA purification system

Working with DNA
Sterile Technique

Biochemical Protocol 2. Microscope Technology (1)

3D-rendering

Additional Applications of Confocal Microscopy

Background/Colophone

Comparison of Electron Micrographs

Confocal microscope technique

Confocal Microscopy (Information about the history, confocal theory, principles and optics as well as a selection of general links)

Conventional VS Laser scanning Microscopy Techniques

Creatingand enhancing professional graphics using Adobe PhotoshopCS3

Dark field illumination, Larry Legg

Digital Image Ethics (A short history of some of the issues that journalists have struggled with and ten suggested guidelines for working with digital images)

Digital Microscope Camera Technology
Confocal Laser Scanning Microscopy
Light Microscope Techniques
Materials, Forensics, Geological Microscopy
Electron and Light Microscopy Web Resources

Educational Resources (On-line resources for K-12 teachers & students)

Electron Microscope Techniques
Historical and Antique Microscopes
Scanning Probe and Atomic Force
Energy Dispersive Microanalysis EDX

Electron microscopy

Electron Microscopy (Information about history, principles, sample preparation techniques and www resources for TEM & SEM)

Electron microscopy

Facilities

Fluorescence Techniques (Links with information about fluorescent dyes, antibodies and resources for sample preparation)

Free Publications (Seven publications for professionals that cover microscopy & imaging from a variety of perspectives)

Hardware (Information about digital cameras, scanners and printers)

Histology (Links to sample preparation protocols and staining techniques, immuno/histo/cyto-chemistry, collections of images of tissues prepared for histology and safety)

Biochemical Protocol 2. Microscope Technology (2)

Image capture
Image analysis

Image Galleries (Links to collections of beautiful images that use a number of microscopic imaging techniques0

Introduction

Introduction (A look at the basics of what makes up a digital image, color and file formats, as well as a few links on human vision)

Laser microscopy

Laser microscopy: stereo-viewer

Light & Microscopes Larry Legg's funky introduction

Light Microscopy (Links to excellent resources that introduce concepts of light & optics, as well as how light microscopes work)

Light microscopy

McCrone Research Institute Teaching light microscopy, pollen and spore identification

Measurement of Cell Organelles

Measurements: Ocular and Stage Micrometers

Measuring Area

Measuring Depth

Measuring Volume

Microscopy - Wikipedia, the free encyclopedia

Microscopy Resources (General information about professional societies, vendors, web resources and email discussion groups)

Microscopy-UK general resource, training, and supplies

Phase Contrast illumination and other methods, Larry Legg

Polarized light illumination - Do-it-yourself, cheap

Polarized Light illumination, David Walker

Preparation in TEM

Sample Preparation of Confocal Microscopy

Software (Links to resources for several commercial and freeware image editing programs)

Stereo vision

The Bright Field Microscope

The Inverted Phase Microscope

The Phase Contrast Microscope

The Transmission Electron Microscope

Use of Darkfield Illumination

Use of Oil Immersion (100x)

Video microscopy (Microscopy & Imaging Resources on the WWW)

Biochemical Protocol 3. Histochemistry Protocols

Alkaline Phosphatase Localization

Autoradiography

Basophilia

Cell fixation - room temperature paraformaldehyde (good for paraspeckles/speckles for 5 min; most other things for 10 min)
Cell fixation - methanol

Conjugating antibodies to fluorescent probes

Histology Protocol

Immunoflourescence of MEFs
Mouse Necropsy
Necropsy form
Paraffin embedding and Sectioning
Staining Protocols
H & E staining
BrdU Staining Histological Sections
X-gal Staining
HRP-DAB Immunostaining
AP-BCIP/NBT Immunostaining
GFP & BrdU In Situ Immunoflourescence
Fixation, permeabilization and immunostaining of cells
(contains images and movies)
Cell fixation - warm paraformaldehyde (preserves microtubules)

Immunofluorescent Localization of Tubulin

Methyl Green-Pyronin Staining of DNA and RNA

Periodic Acid Schiff (PAS) Reaction

Selective Staining: Prepared Slides

The Feulgen Procedure for DNA

Biochemical Protocol 4. Protein

Affinity column preparation and Immunoprecipitation

Affinity purification of Antibodies

B-PER Bacterial Protein extraction reagent (Pierce)

Bradford quantification of protein concentration

Crosslinking Protocol

Crosslinking the antibody to the beads protocol | Abcam

Factor Xa Cleavage of GST fusions

GST-fusion protein injection into rabbits

GST-protein fusion expression, purification and other uses

His-fusion protein purification (Novagen/Qiagen)

Histone Kinase Assay
Western blot
p27 ELISA

Immunoblot Anaylsis

Immunoprecipitation

Protein Interactions Captured by Chemical Cross-linking: One-Step Cross Linking.

Protein precipitations

Protein preparation (from plants tissue)

Protein staining

Protocol: Cross-linking of IgG to Protein A or G Beads: NEB

Radio-labeling of protoplasts

SDS/Native PAGE

Silver staining

Stripping blots to reprobe with different antibodies

UV Crosslinking Protocol

Western blots

Biochemical Protocol 5. Cell Fractionation

Cell Fractionation Protocol.
Chromatin isolation
Spliceosome purification

Cell Fractionation Protocol.
Covalently conjugate antibodies to beads for affinity purification
Nucleolar isolation
(contains images and movies)
Cajal body isolation
(contains images and movies)
Lysing cells for release of both cytoplasmic and nuclear proteins

Equilibrium Density Gradient - Percoll

Low Speed Separaton of Cells

Preparation of nuclear extract from plated cells

Sedimentation Densities

Separation of Cells by Velocity Sedimentation

Sucrose Fractionation of Castor Bean