As well as the IIS guest user pay attention to the user used by the Application Group. Your ASP.Net code could be running as either, I’m not sure what the rules are but it might be the thing that you are overlooking when having permission problems.

Also remember to make sure that ASP or ASP.Net is enabled if you are running Windows Server 2003. It’s another source of disappointment when setting a up a new site on new boxes.