Prediction of transmembrane alpha-helices and signal peptides RET4 HUMAN

TMHMM

First, we predicted the transmembrane helices with TMHMM.

Figuer 1: Prediction of TMHMM for the transmembrane helices of RET4_HUMAN

start position	end position	location
1	201	outside

TMHMM predicts no transmembrane helices (compare Figure 1). The whole protein is located at the extracellular space.

Comparison with the real structure of the protein:

Figure 2: Comparison between real occurring transmembrane helices and the TMHMM result.

The TMHMM prediction is completely right, as we can seen on Figure 2. Therefore, you can see TMHMM can also predict, that a protein is not a transmembrane protein.

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Phobius and PolyPhobius

The next tools we used for the prediction are Phobius and PolyPhobius.

Figure 3: Prediction of Phobius for the transmembrane helices and signal peptides of RET4_HUMAN

Figure 4: Prediction of PolyPhobius for the transmembrane helices and signal peptides of RET4_HUMAN

Phobius			PolyPhobius
start position	end position	prediction	start position	end position	prediction
Signal peptide prediction
1	2	N-Region	1	3	N-Region
3	13	H-Region	4	13	H-Region
14	18	C-Region	14	18	C-Region
Summary signal peptide
1	18	secretory signal peptide	1	18	secretoy signal peptide
Transmembrane helices prediction
19	201	outside	19	201	outside

Both methods predict a signal peptide for the secretory pathway (compare Figure 3 and Figure 4). This result is correct.

Comparison with the real structure of the protein:

Figure 5: Comparison between the prediction of Phobius and the real protein

Figure 6: Comparison between the prediction of PolyPhobius and the real protein

Both methods show exactly the same result, as can be seen on Figure 5 and Figure 6.

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OCTOPUS and SPOCTOPUS

Next, we used OCTOPUS and SPOCTOPUS to predict the transmembrane helices and also the signal peptide.

Figure 7: Prediction of OCTOPUS for the transmembrane helices of RET4_HUMAN

Figure 8: Prediction of SPOCTOPUS for the transmembrane helices of RET4_HUMAN

OCTOPUS			SPOCTOPUS
start position	end position	prediction	start position	end position	prediction
1	1	inside	1	5	N-terminal of a signal peptide
2	23	TM helix	6	19	signal peptide
24	201	outside	20	201	outside

As before by HEXA_HUMAN, OCTOPUS predicts a transmembrane helix (Figure 7), whereas SPOCTOPUS predicts the signal peptide (Figure 8).

Comparison with the real structure of the protein:

Figure 9: Comparison between the prediction of OCTOPUS and the real protein

Figure 10: Comparison between the prediction of SPOCTOPUS and the real protein

Both methods show exactly the same prediction result, except the beginning of the protein. As we can see on Figure 9 and Figure 10, these results agree totally with the real occurring structure.

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TargetP

All of our proteins are proteins from human and archaea, so therefore we only use the non-plant option of TargetP.

Location	Probability
mitochondrial targeting SP	0.242
secretory pathway SP	0.928
other	0.020

TargetP predicts a secretory pathway signal peptide for this protein, which is completely correct.

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SignalP

For our analysis we used the Hidden Markov Model based and also the neuronal network based prediction.
The prediction with the Hidden Markov Model used three different scores. The S-score which is the score for the signal peptide, the C-score which is the score for the cleavage site and the Y-score which is a combination of the S-score and the C-score and is used to predict the cleavage site, because the Y-score is more precise than the C-score.

Result of the neuronal network

Signal peptide		Cleavage site
start position	end position	start position	end position	prediction
1	18	18	19	signal peptide

Result of the hidden markov model

prediction	signal peptide probability	signal anchor probability	cleavage site start	cleavage site end
signal peptide	1.000	0.000	18	19

Figure 11: Result of the SignalP method based on the neuronal network for RET4_HUMAN

Figure 12: Result of the SignalP method based on the Hidden Markov Model for RET4_HUMAN

Both methods predict a signal peptide for RET4_HUMAN (compare Figure 11 and Figure 12), which is correct.

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