pH: All enzymes have an optimum pH value which is very specific. This is because hydrogen ions and hydroxide ions have very significant changes on the tertiary structure of an enzyme, which can cause substrate to be unable to bind to it if the pH is not the correct value.
Temperature: As enzymes become hotter, they vibrate more. This puts strain on bonds such as hydrogen and ionic bonds. This can cause the bonds to break. If the bonds break then more and more bonds are broken. This will alter the shape of the active site and therefore cause rate of reaction to decrease. If enough bonds break then the tertiary structure will unravel. This is irreversible and the enzyme will be described as being denatured.
Substrate Concentration: Initially rate of reaction increases as substrate concentration increases, as it increases the amount of collisions between enzyme and substrate molecules, thus increasing the amount of enzyme-substrate complexes. At a point, the rate of reaction tails off (plateaus) as all of the active sites are taken up. This means that further substrate concentration increase will have no effect
Enzyme Concentration: As the enzyme concentration increases more active sites become available. This means that more enzyme-substrate complexes form so the rate of reaction will increase. If the enzyme concentration continues to increase then there will be a point where all of the substrate molecules are occupying enzyme active sites. This means that substrate concentration is the limiting factor. Rate of reaction can then only be increased by increasing substrate concentration.